JP7343742B2 - Analyzer and method - Google Patents

Description

The present invention relates to an analysis device and an analysis method for analyzing a sample based on scattered light obtained by irradiating the sample with light.

Traditionally, the size and shape of fine particles in a sample liquid, as well as the size and structure of their aggregates, particularly the condition of nanometer to submicrometer-sized particles and aggregates, have been subject to analysis and evaluation. ing. For example, the state of nano to submicron metal, plastic, and inorganic particles, which are dispersed in a sample liquid and have relatively small interaction with solvent molecules, and their aggregates are analyzed and evaluated. In addition, for example, the state of molecules that strongly interact with and dissolve in solvent molecules, such as small and medium molecules such as peptides and nucleic acids, and biopolymers such as proteins, and their aggregates in the liquid is analyzed and evaluated. is subject to.

In the above analytical evaluation, two light scattering measurement methods are generally used. One of the light scattering measurement methods is dynamic light scattering (DLS), which detects light scattering from an object to be analyzed and evaluated at a predetermined scattering angle and measures the temporal fluctuations (fluctuations) of light scattering. scattering) measurement method. The DLS measurement method mainly provides information regarding the motion of the scatterer to be analyzed.

Another light scattering measurement method is a method of measuring the dependence of the scattering angle on the average intensity of light scattering from an analysis target, and is a static light scattering (SLS) measurement method. The SLS measurement method mainly provides information regarding the structure of the scatterer to be analyzed.

A sample analyzer using the above DLS measurement method or SLS measurement method is described in Patent Document 1, Patent Document 2, and Patent Document 3 described below.

Patent No. 5883233 Patent No. 6757964 Special table 2019-536997 publication

T. Wakamatsu, T. Onoda, M. Makoto, “Time-resolved forward-light-scattering monitoring of protein-lysozyme aggregation in pre-crystalline solutions”, Japanese Journal of Applied Physics, (2018), vol. 57,058003-1 to 058003-3.

The state of the measurement target, which is a sample, changes over time. Sample aging is unavoidable. It is very important to obtain a lot of information on the same measurement target and over the same elapsed time. For example, if DLS and SLS measurements can be performed simultaneously on the same measurement object, very valuable information about the measurement object can be obtained. Although this is just one example, valuable information can be obtained for analyzing and evaluating knowledge about the motion and structure of the target of analysis, such as particles and aggregates.

Although we conducted a search for known literature, we could not find any known literature that describes or suggests the importance of simultaneously measuring DLS and SLS on the same measurement target. Moreover, no known literature suggesting a configuration for realizing this was found.

An object of the present invention is to provide an analysis device and an analysis method that are capable of performing measurements using both the DLS measurement method and the SLS measurement method on the same measurement target at the same elapsed time. .

[First invention]
The first invention to solve the above problem is:
a light source that generates light used for measuring the sample;
a light irradiation unit that irradiates the sample with irradiation light that forms a focal point within the sample based on the light generated by the light source;
a parallel scattered light conversion unit that receives forward or backward small angle scattered light generated in the analysis micro region where the focal point of the irradiation light is formed, and generates parallel small angle scattered light;
a scattered light detection unit for measuring the state of the small angle scattered light from the sample based on the parallel small angle scattered light;
an analysis section including a storage device that analyzes the sample based on the measurement results of the scattered light detection section;
The analysis unit repeatedly stores the measurement results of the scattered light detection unit in the storage device,
The analysis unit calculates a dynamic scattered light component or a static scattered light component, or the dynamic scattered light component and the static scattered light component, based on the measurement results of the scattered light detection unit stored in the storage device. The analyzer is characterized in that it analyzes both components and performs an analysis related to the analysis microregion of the sample.

[Second invention]
A second invention for solving the above problem is, in the analysis device of the first invention,
The scattered light detection unit includes a photodetector,
A microscopic image, which is an enlarged image of the state of the sample present in the analysis microregion, is generated on the photodetector;
The analysis device is characterized in that the analysis section repeatedly captures the microscope image as the measurement result of the scattered light detection section and stores it in the storage device.

[Third invention]
A third invention for solving the above problem is, in the analysis device of the second invention,
The analysis unit stores a moving image of the microscope image in the storage device by repeatedly capturing the microscope image from the scattered light detection unit,
The analysis device is characterized in that the analysis unit analyzes the dynamic scattered light component and the static scattered light component based on the stored moving image.

[Operations and effects of the first to third inventions]
Next, the effects of the first to third inventions will be explained. In the configurations of the first to third inventions, the analysis section has a configuration for repeatedly storing the measurement results of the scattered light detection section in the storage device, for example, a predetermined number of times or a predetermined time. are doing. Therefore, it is possible to instantly obtain the measurement results of the sample required for analysis of dynamic scattered light and static scattered light. The state of a sample often changes over time, but in the first to third inventions, the state of the sample changes over time, and the measurement results of the sample necessary for analysis of dynamic scattered light and static scattered light can be seen to be obtained simultaneously and instantaneously. That is, it is possible to perform analysis and evaluation using the dynamic scatterometry method and the static scatterometry method on the same measurement target area and in the same state over time.

Furthermore, with conventional techniques, the analyzable area within a sample is large, making analysis narrowed down to a minute area impossible. The first invention includes a light irradiation unit that irradiates the sample with irradiation light that forms a focal point within the sample, and the area to be analyzed can be narrowed down to a very small analysis micro area.

For example, the technology described in Patent Document 1 is a measurement and analysis device that acquires relaxation data of a time-averaged autocorrelation function-correlation function, that is, a DLS measurement and analysis device. DLS measurements are performed for each scattering angle by changing the angle. For example, see paragraph [0045] of Patent Document 1.

Furthermore, in Patent Document 1, as another embodiment of a scanning microscopic light scattering measurement and analysis device, an objective lens and a cover member are used to detect scattered light from a sample at a selected scattering angle. Regarding the DLS measurement method for selective detection (for example, as described in paragraph [0103] of Patent Document 1), a scattering angle selection filter was further placed between the objective lens and the imaging lens, and the selection was made in the same manner. A DLS measurement method that selectively detects only scattered light at a scattering angle is disclosed (for example, as described in paragraph [0115] of Patent Document 1).

However, there is no disclosure of a means for measuring scattered light by continuously changing the scattering angle using the cover member and the scattering angle selection filter (herein referred to as a scattering angle selection device for convenience), Using the scattering angle selection device described above, the scattering angle can be changed instantaneously and continuously (for example, in the scattering angle range of 1° to 15°, the angular resolution is about 0.1° or less, and the scattering angle is changed instantaneously in 0.1 seconds or less. SLS measurement, which measures the intensity spatial pattern of scattered light (scattered light intensity distribution with respect to scattering angle), is technically difficult. Therefore, with the technique of Patent Document 1, it is difficult to measure DLS and SLS simultaneously.

Further, Patent Document 1 discloses DLS measurement at multiple points on a sample, for example, DLS measurement on a minute volume while changing measurement positions at intervals of several μm using a scanning mechanism. For example, see paragraph [0007] of Patent Document 1. However, as mentioned above, it is difficult to instantaneously measure SLS from a sample on a microscopic scale by using the scattering angle selection device, and furthermore, it is difficult to simultaneously and instantaneously measure DLS and SLS on a microscopic scale. there were.

In addition, in Patent Document 2, in the measurement of forward small-angle scattered light or backward small-angle scattered light from a protein crystallization solution, these scattered lights are captured in a predetermined scattering angle range, converted to parallel scattered light, and the scattered light is The intensity distribution (distribution function for scattering angle) was measured all at once in real time. For example, see paragraph [0070] of Patent Document 2.

Patent Document 2 discloses a real-time SLS measurement method and a method for analyzing the SLS measurement data. For example, see paragraph [0107] of Patent Document 2. However, a real-time DLS measurement method and its analysis method are neither disclosed nor suggested, and furthermore, a simultaneous DLS and SLS measurement method and its analysis method are not described. Furthermore, in Patent Document 2, the incident light that enters the solution cell is parallel light (collimated light). For example, see paragraph [0056] of Patent Document 2. The size of the incident light beam is, for example, about 0.7 mm in diameter. For example, see paragraph [0133] of Patent Document 2. For this reason, it is difficult to carry out simultaneous measurement of DLS and SLS on a microscopic scale in a microscopic area of a sample, for example, on the order of several tens of micrometers to one hundred micrometers or less, and to locally analyze and evaluate particles and aggregates. Therefore, in Patent Document 2, DLS and SLS measurements are performed simultaneously and instantaneously, and the diffusion coefficient and size of particles and aggregates to be analyzed, as well as the structure of aggregates, etc., are analyzed, and the solution structure, etc., is analyzed on a microscopic scale. It is difficult to analyze and evaluate the microstructure.

Furthermore, the technique described in Patent Document 3 generates an interference beam in the sample using an optical element including an axicon lens, forms a dark region that is not substantially irradiated, and eliminates scattered light in the dark region in the far field. It is a technique that performs DLS and/or SLS measurements at low scattering angles of 0° to 10° from the irradiation axis. A plurality of the scattered light detectors are arranged in the range of 0° to 10°. For example, see paragraph [0015] of Patent Document 3. For example, the detector or detectors are configured to detect scattered light in a range of 1° or 2° or 3° or 4° or 5°. For example, see paragraph [0016] of Patent Document 3.

That is, since the scattered light is detected at discrete scattering angles using a plurality of detectors, it is difficult to instantaneously measure a continuous SLS spatial pattern. Here, it is stated that when measuring scattered light at any angle between 0° and 10°, an optical fiber is coupled to the detector. For example, see paragraph [0016] of Patent Document 3. However, there is no disclosure of a specific method; instead, the scattered light taken in by the optical fiber at various angles is detected by a detector and converted into an SLS signal, that is, a scattered light intensity function (the scattering angle dependence of the scattered light intensity). It is technically difficult to obtain. Therefore, in Patent Document 2, it is difficult to perform DLS and SLS simultaneously and instantaneously.

[Operations and effects of the second invention and the third invention]
In addition to the above description, in a second invention or a third invention, the method receives forward or backward small-angle scattered light from an analysis microregion in which the focal point of the irradiation light is formed in the sample, and receives parallel scattered light. It has a configuration in which the parallel small-angle scattered light formed by the converter is detected by the scattered light detector. In the second aspect of the invention, as described above, a microscopic image is generated on the photodetector, which is an enlarged image on a microscopic scale of the state of the sample existing in the microscopic analysis area. Furthermore, by continuously and repeatedly capturing these microscopic images, it has become possible to measure the changes that occur moment by moment in the microscopic images as moving images. DLS analysis can be performed by analyzing moment-to-moment changes in a specific region. Furthermore, SLS analysis can be performed by capturing and analyzing the microscope image in a region extending in the radial direction from the optical axis. In other words, the process of change in a sample over time that occurs in the same area can be captured as a moving image of a microscope image, which is an image enlarged on a microscope scale. Further, from this moving image, it is possible to obtain analysis results of both DLS analysis and SLS analysis at a specific time point in the process of change of the sample over time. Moreover, analysis can be performed using the measurement results enlarged on a microscopic scale. In the conventional technology, such an analysis could not be performed.

[Fourth invention]
A fourth invention for solving the above-mentioned problem is an analyzer according to one of the first to third inventions, comprising:
A scattering angle filter and a scattered light concentrator are provided between the parallel scattered light converter and the scattered light detector, and the scattering angle filter divides the small angle scattered light from the parallel scattered light converter. , the arc-shaped specific angle scattered light forming an arc shape in a plane perpendicular to the optical axis of the parallel small-angle scattered light passes through the scattered light condenser, and is detected by the scattered light detection. incident on the photodetector of the
An analysis device characterized in that the microscope image, which is an enlarged display of the state of the sample within the analysis microregion, is detected by the photodetector by the incidence of the parallel small-angle scattered light on the photodetector. It is.

[Fifth invention]
A fifth invention for solving the above problem is an analysis device according to the fourth invention, which includes:
The analysis unit repeatedly captures and stores the microscopic image, which is an enlarged display of the state of the sample within the analysis microregion detected by the photodetector, thereby determining the state of the sample within the analysis microregion. The analysis device is characterized in that the moving image of the microscope image showing the state in an enlarged manner is stored in the storage device.

[Sixth invention]
A sixth invention for solving the above problem is an analysis device according to one of the fourth invention or the fifth invention, comprising:
The scattering angle filter is made of a material that blocks light, and has an arc-shaped aperture whose radius is a length corresponding to the specific angle with respect to the optical axis of the parallel small-angle scattered light. This is an analytical device with special features.

[Operations and effects related to the fourth to sixth inventions]
As will be explained below based on the embodiments shown in FIGS. 10 to 14, small-angle scattered light 160 is generated by the sample existing in the analysis micro region 156 within the sample 154 based on the irradiation light 148 irradiated from the light irradiation unit 140. Made. This small-angle scattered light 160 is converted into parallel small-angle scattered light 168 by a parallel scattered light converter 170. When the parallel small-angle scattered light 168 enters the scattering angle filter 180, the arc-shaped specific angle scattered light 186 having a specific scattering angle and having an arc shape in a plane perpendicular to the optical axis 146 passes through the scattering angle filter 180. The light passes through and enters the scattered light condensing section 190. When this circular arc-shaped specific angle scattered light 186 enters the photodetector 202 via the scattered light condenser 190, an image representing the state of the sample existing in the analysis microregion 156 is converted into an enlarged microscope image. , detected by the photodetector 202. This microscope image can be taken into the analysis unit 250 and stored as a detection result of the photodetector 202.

Although we searched for known literature, we could not find any known example disclosing or suggesting a technique capable of displaying an enlarged image representing the state of a sample present in the analysis microregion 156, that is, a microscopic image. Note that Patent Document 1 describes a scattering angle selection filter 19. However, the aperture of this scattering angle selection filter 19 is a point and does not have an arc-shaped spread. For this reason, the microscopic image cannot be created.

[Seventh invention]
A seventh invention for solving the above problem is an analysis device according to one of the first to sixth inventions, comprising:
The analyzer is characterized in that the incident light that enters the light irradiation section is a light beam having a circular cross-sectional shape, and the diameter of the light beam of the incident light is 1.6 mm or less.

[Eighth invention]
An eighth invention for solving the above problem is an analysis device according to one of the first to seventh inventions, comprising:
The light irradiation unit includes a condensing lens for condensing light and a lens adjustment mechanism for moving the condensing lens along an optical axis of the incident light incident on the light irradiation unit. ,
The lens adjustment mechanism is an analysis device characterized in that a distance over which the condensing lens can be moved along the optical axis is ±10 mm or less, and a minimum length over which the movement can be adjusted is 50 μm or less.

[Ninth invention]
A ninth invention for solving the above problem is an analysis device according to one of the first to eighth inventions, comprising:
An analysis device characterized in that the diameter of a minimum beam spot formed inside the sample by irradiating the sample with the irradiation light from the light irradiation section is in the range of 10 μm or more and 100 μm or less. It is.

[10th invention]
A tenth invention for solving the above-mentioned problem is an analysis device according to one of the first to ninth inventions, comprising:
It has a sample part,
The sample section includes a sample cell for holding the sample and a sample position adjustment mechanism for adjusting the position of the sample cell,
The analysis unit outputs an adjustment image for adjusting the position of the sample cell by the sample position adjustment mechanism based on the measurement result of the scattered light detection unit,
The analysis unit repeatedly stores the measurement results of the scattered light detection unit in the storage device based on measurement instructions for performing the analysis,
The analysis device is characterized in that the analysis section performs the analysis based on the measurement results of the scattered light detection section that are repeatedly stored in the storage device.

[Operations and effects of the tenth invention]
The effects of the tenth invention will be explained. In the first invention, by forming the focus of the irradiation light on the region to be analyzed within the sample, the state of the region to be analyzed can be accurately measured. However, in order to take advantage of this effect, it is necessary to adjust the position of the sample cell, for example in a plane perpendicular to the optical axis, using a sample position adjustment mechanism so that the focus of the irradiation light is formed on the area to be analyzed. becomes. In the present invention, an adjustment image is displayed so that this work can be performed easily and accurately. This adjustment image is drawn based on, for example, small-angle scattered light generated from the sample at the current focal position of the irradiation light. By adjusting the position of the sample cell using the sample position adjustment mechanism while viewing this image, the position of the sample cell can be adjusted more accurately and more smoothly.

[Eleventh invention]
An eleventh invention for solving the above problem is, in the analyzer according to one of the first to tenth inventions, a maximum incidence of irradiation light irradiated from the light irradiation part to the sample with respect to an optical axis. The analyzer is characterized in that the angle θi is in the range of 0.5 degrees to 20 degrees.

[12th invention]
A twelfth invention for solving the above-mentioned problems is an analysis device according to one of the first to eleventh inventions, comprising:
a first step in which the analysis unit repeatedly captures the measurement results of the scattered light detection unit based on set sampling conditions;
a second step of extracting a measurement result of a dynamic scattered light analysis region and a measurement result of a static scattered light analysis region from the measurement results of the scattered light detection unit that have been repeatedly captured;
performing calculations for dynamic scattered light analysis based on the measurement results of the dynamic scattered light analysis region and calculations for static scattered light analysis based on the measurement results of the static scattered light analysis region; 3 steps and
This is an analysis method characterized by having the following.

[13th invention]
A thirteenth invention for solving the above problem is, in the analysis method of the twelfth invention,
After the first step of repeatedly capturing measurement results from the scattered light detection unit based on the set sampling conditions, the analysis unit stops execution of the first step for a set imaging interval,
The analysis unit starts executing the first step again after the set imaging interval ends,
The analysis unit calculates the first step according to a set number of times the first step is executed or a set total imaging time that is a time period in which execution of the first step and subsequent stopping of execution are repeated. An analysis method comprising repeating the steps of executing the step and stopping the execution of the first step during the imaging interval.

[14th invention]
A fourteenth invention for solving the above problem is, in the analysis method of one of the twelfth or thirteenth inventions,
The analysis unit operates in an adjustment mode and a measurement analysis mode based on instructions,
In the operation in the adjustment mode, the third step of performing the calculation for the dynamic scattered light analysis and the calculation for the static scattered light analysis based on the measurement data from the scattered light detector is stopped. This is a characteristic analysis method.

According to the present invention, it is possible to obtain an analysis device and an analysis method that can perform both the DLS measurement method and the SLS measurement method on the same measurement target at the same elapsed time.

FIG. 2 is an explanatory diagram for explaining an embodiment of an analysis device that analyzes a microscopic region of a sample to which the present invention is applied. FIG. 2 is an explanatory diagram illustrating the configuration and operation of a light irradiation section, a sample section, and a parallel scattered light conversion section in the configuration shown in FIG. 1; FIG. 2 is an explanatory diagram illustrating the relationship between irradiation light and generation of small-angle scattered light in an analysis micro region of the sample shown in FIG. 1. FIG. It is an explanatory view of a microscope image measured in an example to which the present invention is applied. FIG. 2 is an explanatory diagram for explaining an analysis method based on a microscopic image of small-angle scattered light. FIG. 3 is an explanatory diagram illustrating a detection surface of an example of a photodetector in a scattered light detection section. FIG. 3 is an explanatory diagram illustrating a relationship between a detection result detected by a scattered light detection unit and a storage state. FIG. 3 is an explanatory diagram illustrating the relationship between a microscope image based on small-angle scattered light detected by a scattered light detection unit and a DLS analysis region R1 and an SLS analysis region R2. FIG. 6 is an explanatory diagram illustrating the relationship between the SLS analysis region R2 and the detection results in the detection area of the scattered light detection unit. It is an explanatory view explaining other examples to which the present invention is applied. 11 is an explanatory diagram illustrating the operation of a scattering angle filter and a scattered light condensing section in another embodiment shown in FIG. 10. FIG. It is an explanatory view explaining a scattering angle filter. It is an explanatory view when a camera with a lens is used as a scattered light detection section. This is another embodiment in which a camera with a lens is used as a scattered light detection unit to measure a scattered light image. FIG. 15 is an explanatory diagram illustrating an example of the operation of the analyzer. 16 is a flowchart for performing the operation of FIG. 15. FIG. 2 is an explanatory diagram illustrating a storage state of measurement results in a storage device and a storage state of analysis results using the stored data.

1. Description of the analyzer 100 for analyzing the sample 154 based on scattered light 1.1 Introduction In this specification, components given the same reference numerals perform the same operations and effects, and produce the same effects. In order to avoid complexity, repeated explanations of structures with the same reference numerals may be omitted. Furthermore, in the description of this specification, the terms "detection", "detection", "measurement", and "measurement" are not strictly distinguished. They are described as having the same meaning. Furthermore, there is no strict distinction between the terms "irradiate" and "incident". They are described as having the same meaning.

1.2 Overview of Analyzer 100 FIG. 1 shows the configuration of an analyzer 100, which is an embodiment to which the present invention is applied. As will be described in more detail below, a light source 110 generates light 112 suitable for illuminating a sample 154 to be analyzed. Light 112 generated by the light source 110 is guided to an incident light adjusting section 120, which adjusts the light 112 to generate incident light 126. Incident light 126 is introduced into light irradiation section 140 . The light irradiation unit 140 generates irradiation light 148 for irradiating the sample 154 based on the introduced incident light 126. The irradiation light 148 is irradiated onto the analysis microregion 156 of the sample 154 and forms a focus on the analysis microregion 156 . By irradiating the irradiation light 148, scattered light is generated from the analysis micro region 156 of the sample 154 on the front side and the rear side of the sample 154. Of the front side and rear side scattered light, at least one small angle scattered light 160 is converted into parallel small angle scattered light 168 by the parallel scattered light converter 170, and detected by the scattered light detector 200. Based on the detection result of the scattered light detection section 200, the analysis section 250 performs analysis on the sample 154, and the analysis result is output from the input/output device 266 of the analysis section 250.

Note that the small-angle scattered light 160 generated on the rear side, the configuration for converting the small-angle scattered light 160 generated on the rear side into parallel small-angle scattered light 168, and the detection of the parallel small-angle scattered light 168 based on the small-angle scattered light 68 on the rear side The configuration of the scattered light detection section 200 is approximately the same as that in which the small-angle scattered light 160 generated on the front side is converted into parallel small-angle scattered light 168 by the parallel scattered light conversion section 170 and detected by the scattered light detection section 200. Since they have the same configuration and substantially the same effects, descriptions using figures will be omitted.

The irradiation light 148 generated by the light irradiation section 140 is very important for improving the analysis accuracy of the sample 154. In the light irradiation section 140, the irradiation light 148 is formed based on the incident light 126 generated by the incident light adjustment section 120, so the incident light adjustment section 120 generates the incident light in a state suitable for generating the irradiation light 148. It is important to generate 126. Further, it is desirable to prevent stray light other than the incident light 126 from entering the light irradiation unit 140. The optical aperture 138 is provided for this purpose. Furthermore, the optical aperture 138 has the effect of preventing the reflected light from the sample section 150, the parallel scattered light conversion section 170, etc. from entering the sample 154 of the sample section 150 again.

In the embodiment shown in FIG. 1, a configuration is described in which the small-angle scattered light 160 generated in the forward direction out of the small-angle scattered light 160 generated by the sample 154 based on the irradiation light 148 is used. Furthermore, in the description of FIG. 1, only the small-angle scattered light 160 generated on one side with respect to the optical axis 146 is described. However, this is done to avoid the complexity of explanation and illustration. Scattered light is generated from the sample 154 not only in the forward direction but also in the backward direction. Regarding the small-angle scattered light 160 of this scattered light, small-angle scattered light 160 generated toward the rear as well as the front can be similarly utilized. Furthermore, the small-angle scattered light 160 that is generated forward and backward is generated not only at a specific angle with respect to the optical axis 146 but also over the entire circumference. It is possible to utilize the scattered light generated over the entire circumference. In FIG. 1, small-angle scattered light 160 limited to only a specific angle of the scattered light is shown to be incident on the scattered light detection section 200. However, this is described as an example, and scattered light is generated over the entire circumference of the optical axis 146, including portions other than the small-angle scattered light 160 shown in FIG. It can be input to the scattered light detection section 200 and detected as necessary.

In the analyzer 100, it is preferable to have a degree of freedom in the arrangement relationship between the light irradiation section 140 and the incident light adjustment section 120, and when it is necessary to change the optical axis of the incident light 126 with respect to the light irradiation section 140. occurs. In this case, by providing the optical path changing section 130, it is possible to change the optical axis. In this embodiment, the optical path changing section 130 is provided between the incident light adjusting section 120 and the optical diaphragm 138, but this is just an example, and it is also possible to provide it in other parts as necessary. It may be provided.

As described above, the light irradiation section 140 converts the incident light 126 from the optical path changing section 130 into a focused beam having a focal point, and irradiates the sample 154 as irradiation light 148 . The condensing lens 142 of the light irradiation unit 140 is set so that the focus of the irradiation light 148 coincides with the position of the target region to be analyzed on the sample 154, that is, the focus is located on the analysis microregion 156 to be analyzed. has been done. In other words, the positional relationship between the condenser lens 142 and the sample 154 is set based on the focal length of the condenser lens 142.

The sample 154 is housed by a sample position adjustment mechanism 158 provided in the sample section 150 so that the focal position of the irradiation light 148 is at the analysis microregion 156, which is the point to be analyzed inside the sample 154. It is possible to finely adjust the position of the sample cell 152 in three dimensions. With such a configuration, as will be described in detail below, by adjusting the sample position adjustment mechanism 158 so that the irradiation light 148 is focused on the analysis microregion 156, which is the part of the sample 154 that is desired to be analyzed, Small-angle scattered light 160 can be accurately generated from the region to be analyzed. In this embodiment, since the small-angle scattered light 160 generated at a target portion within the sample 154 can be used, an analysis result of the target portion can be obtained.

The relationship between the focal position of the irradiation light 148 and the sample 154 in the optical axis 146 direction is determined by the focal length of the condenser lens 142, so the distance between the condenser lens 142 and the sample cell 152 on the optical axis 146 is By accurately maintaining the distance between lens 142 and sample cell 152, the focus of illumination light 148 can be located inside sample 154. However, where on the surface of the sample 154 perpendicular to the optical axis 146 the irradiation light 148 is focused varies depending on the operator's intention. For this reason, it is important that the sample position adjustment mechanism 158 can move the sample cell 152 in a plane perpendicular to the optical axis 146. As will be explained again below, in this embodiment and other embodiments explained below, before starting formal measurement, based on the detection results of the scattered light detection unit 200, the parallel small-angle scattered light 168 is The adjustment image is displayed by the analysis unit 250 on the input/output device 266 as an adjustment image. While viewing this adjustment image, the sample position adjustment mechanism 158 moves and adjusts at least the sample cell 152 in a plane perpendicular to the optical axis 146. This allows the irradiation light 148 to be focused on the region desired to be analyzed. The focal region of the irradiation light 148 becomes a desirable analysis microregion 156.

After the analysis microregion 156 determined by the focal position of the irradiation light 148 is set, the input/output device 266 of the analysis unit 250 issues an instruction to officially start measurement by the operator. Thereby, the analysis unit 250 captures measurement results necessary for analysis of dynamic scattered light and static scattered light from the scattered light detection unit 200. The captured measurement results of the scattered light detection unit 200 are stored in the storage device 290 of the analysis unit 250 in association with the position of the measurement element on the measurement surface of the scattered light detection unit 200. The analysis micro-area 156 to be measured can be accurately set, and the measurement data repeatedly taken from there is stored in association with the positional relationship on the measurement surface of the photodetector 202, which is associated with the angle information with respect to the optical axis 146. Therefore, from the stored data, highly accurate analysis of both dynamic light scattering and static light scattering, which will be explained below, becomes possible. The stored measurement results are values measured for the common analysis microregion 156 and at the same temporal timing.

A portion of the irradiated light 148 irradiated onto the sample 154 becomes scattered light, but most of it becomes transmitted light 164 that penetrates the sample 154. The transmitted light 164 is converted into light parallel to the optical axis 146 by the parallel scattered light converter 170, and is blocked by the transmitted light shield 176 disposed behind or in front of the parallel scattered light converter 170. On the other hand, of the scattered light generated by the analytical micro region 156 of the sample 154, the small-angle scattered light 160, which has an angle close to the transmitted light 164, or in other words, has an angle smaller than the maximum angle θ0S described in FIG. 3, is converted into parallel scattered light. The light is converted into parallel small-angle scattered light 168 by the section 170, and detected by the scattered light detection section 200.

In the embodiment shown in FIG. 1, it is possible to irradiate the irradiation light 148 narrowly to the analysis micro region 156 of the sample 154. Therefore, scattered light generated in the analysis microregion 156 of the sample 154 to be analyzed can be detected in real time by the scattered light detection unit 200 for both dynamic light scattering and static light scattering. The state of particles and aggregates inside can be analyzed in real time by the analysis section 250.

The analysis section 250 detects both dynamic light scattering and static light scattering, analyzes various states of particles and aggregates in the sample 154 based on these detection results, and the results are sent to the analysis section 250. It can be output from the input/output device 266. Information representing various states includes, for example, the diffusion coefficient of particles and aggregates, the size of particles and aggregates, and the structural parameters of particles and aggregates, and these analysis results are sent from the input/output device 266 as analysis results. Output.

In the embodiment shown in FIG. 1, a configuration for detecting the forward small-angle scattered light 160 out of the forward and backward small-angle scattered light 160 generated by the sample 154 is shown as a representative example. As described above, the small analysis region 156 of the sample 154 generates small-angle scattered light 160 not only in the front but also in the rear. Therefore, even if the small-angle scattered light 160 generated not only in the front but also in the rear direction is used, analysis results can be obtained in the same manner as in the case where the small-angle scattered light 160 in the front is used. Therefore, the backward small-angle scattered light 160 can be used instead of the forward small-angle scattered light 160. In the embodiments described below, examples using forward small-angle scattered light will be shown and explained as representative examples of embodiments based on the forward small-angle scattered light 160 or the backward small-angle scattered light 160. The technical operation and effects are essentially the same for both embodiments. Note that the transmitted light 164 exists in the front in the direction of light propagation, and does not exist in the rear. When using the rear small-angle scattered light 160, there is no need to consider the overlap between the transmitted light 164 and the small-angle scattered light 160. However, it is necessary to consider the overlap with the irradiation light 148. It is also necessary to consider the relationship between the light irradiation section 140 and the parallel scattered light conversion section 170.

1.3 Description of each component constituting the analyzer 100 (1) Description of the light source 110 The light source 110 shown in FIG. is required to occur. Based on the light 112 generated by the light source 110, the incident light adjustment unit 120 forms incident light 126, and based on the incident light 126, the light irradiation unit 140 forms irradiation light 148 to irradiate the sample 154. When the sample 154 is irradiated with the irradiation light 148, it is desirable that scattered light, particularly small-angle scattered light 160, be efficiently generated from the sample 154. The light source 110 is required to generate light 112 with properties suitable for efficiently generating small angle scattered light 160 in the sample 154.

Various lasers can be used as the light source 110, such as various gas lasers, semiconductor lasers (LDs), diode pumped solid state (DPSS) lasers, and light emitting diodes (LEDs). Among them, the light source 110 is preferably a high-intensity laser light source that has a long coherence length and oscillates at a single wavelength in a single longitudinal mode. An example of such a laser light source is a DPSS laser. A DPSS laser is a small laser light source that has a built-in optical resonator necessary for laser light oscillation. This DPSS laser has the advantage of being able to provide stable, high-output laser light with low power and high efficiency. Therefore, it is suitable as the light source 110 of the analyzer 100 according to the embodiment of the present invention.

Further, it is preferable that the driving power source and the light source main body of the light source 110 are small and have a size suitable for being incorporated into the analysis device 100 for analyzing particles and aggregates. For this reason, analysis targets include, for example, small and medium molecules such as peptides and nucleic acids, biopolymers such as proteins, and their aggregates, as well as nano to submicron particles such as metals, plastics, and inorganic substances, and their aggregates. In this case, it is preferable to use a laser light source of several mW to several tens of mW.

The power of the light 112 output by the light source 110 is determined by the intensity of the scattered light of the sample 154 to be analyzed, the sensitivity of the photodetector 202 of the scattered light detection unit 200, the sensitivity of the light detection camera 204, and the scattered light intensity measurement. Adjustment in accordance with one or more of the amplification degree of the amplifier used, the exposure time for measuring the intensity of scattered light by the photodetector 202, the exposure time for measuring the intensity of scattered light by the photodetector 202, etc. is suitable. Note that the intensity of the incident light 126 on the sample 154 can be adjusted to an appropriate intensity range by adjusting the intensity of the light 112 from the light source 110 with the light amount adjustment element 122 of the incident light adjustment section 120.

It is preferable that the beam size of the light 112 emitted from the light source 110 is as small and narrow as possible. That is, it is preferable that the light beam be a spot-like light beam. Further, it is preferable that the light 112 from the light source 110 is parallel light, that is, collimated light. Furthermore, it is preferable that the cross section of the light 112 is circular from the viewpoint of being widely applicable to the analysis of various types of samples and achieving high detection accuracy. Regarding the irradiation light 148 from the light irradiation unit 140, it is important that the light can be accurately focused on the analysis microregion 156, which is the analysis part of the sample 154. From this point of view, in this embodiment, the cross-sectional size of the light 112 is That is, it is preferable that the beam size is 1.6 mm or less in diameter. More preferably, the cross-sectional size of the light 112 is 1 mm or less. The cross-sectional size, that is, the beam size, of the incident light 126 that enters the light irradiation unit 140 is adjusted by the incident light adjustment unit 120, but if the beam size of the light 112 is 1.6 mm or less in diameter, particularly 1 mm or less, The beam size of the incident light 126 is determined with almost no adjustment of the beam size in the incident light adjustment unit 120, and it is easy to maintain the beam size of the incident light 126 to a diameter of 1.6 mm or less, and especially to a diameter of 1 mm or less. becomes.

The spread angle of the light 112 emitted from the light source 110 greatly influences the narrowing down accuracy at the focal position of the irradiation light 148 irradiated from the light irradiation section 140. It is very important to make the size of the irradiation light 148 as small as possible at the focal position in order to improve analysis accuracy. From this relationship, it is preferable that the spread angle of the light 112 is 1.5 mrad or less, particularly 1 mrad or less in terms of the total radiation angle. When the spread angle of the light 112 emitted from the light source 110 is large, a small spot-shaped light beam necessary for detection is made to enter the sample 154 using a condensing lens group of the light irradiation unit 140, such as an objective lens. becomes difficult. Furthermore, the relationship between the small-angle scattered light 160 and the scattering angle is disturbed, making it difficult to accurately convert the scattered light from the sample into parallel light within a predetermined radiation scattering angle range by the lens group of the parallel scattered light converter 170. becomes. These causes a decrease in analysis accuracy.

There is no particular restriction on the polarization of the light 112 emitted from the light source 110. Polarized light such as linearly polarized light, circularly polarized light, elliptically polarized light, etc. can be used. It may also be non-polarized light. Among these, linearly polarized light such as TE polarized light and TM polarized light is easy to use.

When using the depolarized light scattering method to analyze various motion elements such as the target particles of the sample 154 or their aggregates, such as translational motion and rotational motion, or when analyzing the orientation of crystal grains in thin films of polymers, etc. When analyzing the polarization, light with a known polarization state, for example, linearly polarized light, is used as the incident light. A scattered light component polarized parallel to the incident light and a scattered light component polarized perpendicular to the incident light are detected separately using a polarizer or the like. For example, when scattered light in the H polarization state, which is the polarization state of horizontally polarized light, is detected in contrast to the V polarization state, which is the polarization state of vertically polarized incident light, it is written as "VH". Ru. Measure “VV” and “VH” as well as “HH” and “HV” and evaluate them to determine the translational motion and rotational motion of the particles to be measured in the liquid, their aggregates, etc. The orientation of crystal grains in thin films of molecules can be analyzed. Furthermore, high-precision scattered light measurement requires stability of the incident light. Therefore, it is preferable that the output stability of the light source 110 is several percent or less per hour. Furthermore, in measuring scattered light, it is also possible to avoid the influence of fluctuations in the incident light by monitoring changes in the intensity of the incident light.

The method of measuring the scattered light from the sample 154, especially the DLS measurement method, includes the homodyne measurement method, which obtains a time correlation function by taking the time autocorrelation of the scattered light itself from the scatterer of the sample 154, and the method of measuring the scattered light from the scatterer. There is a heterodyne measurement method in which a reference light modulated at a certain frequency is mixed with light to obtain a time correlation function. The homodyne measurement method used in DLS measurement has a relatively simple optical system, can perform measurement with a high S/N ratio, and can easily obtain a time correlation function of scattered light. On the other hand, the heterodyne measurement method is often used when the scatterers of the sample 154 are in constant motion, such as in electrophoretic measurements.

In DLS measurement using the homodyne measurement method, a time correlation function can be obtained from temporal fluctuations of the scattered light itself using light whose intensity is constant over time as the light 112 generated by the light source 110. Further, as the light 112 generated by the light source 110, modulated light modulated at a certain frequency may be output. For example, the light 112 emitted from the light source 110 may be modulated at a predetermined frequency using a light modulator such as a light chopper that rotates or vibrates at a certain frequency, or a light modulation element such as a light modulation device. It may also be used as the incident light 126.

On the other hand, in DLS measurement using the heterodyne measurement method, light modulated at a certain predetermined frequency can be used as reference light to be mixed with the scattered light from the scatterer of the sample 154. For example, as the modulated reference light, modulated light from the light source 110 can be used as described above. In this case, the light 112 emitted from the light source 110 is branched into, for example, two optical paths, and the light on one optical path is introduced into the light irradiation section 140 as the incident light 126, and the light 148 is introduced from the light irradiation section 140 into the irradiation light 148. The sample 154 is irradiated as follows. Scattered light from the sample 154 based on the irradiation with this irradiation light 148 is guided to the photodetector 202 of the scattered light detection section 200 via the parallel scattered light conversion section 170. Furthermore, the light on the other optical path is modulated to a certain predetermined frequency by the above-mentioned light modulator or light modulation element, and the modulated light is made to enter the photodetector 202 as it is. The dynamic scattered light from the scatterer of the sample 154 and the modulated reference light are mixed and measured by the photodetector 202 . In this way, a time correlation function of the scattered light with respect to the modulated reference light can be obtained.

(2) Description of the incident light adjustment section 120 The incident light adjustment section 120 has a function of adjusting the state of the irradiation light 148 that is irradiated onto the sample 154 provided in the sample section 150. Specifically, for example, the incident light adjustment unit 120 is composed of a group of optical elements, and adjusts the light amount, polarization state, and light beam in order to make the light 112 from the light source 110 into irradiation light 148 suitable for the sample 154. Adjust the shape, size, etc. The incident light adjusted by the incident light adjustment unit 120 has its optical path changed by the optical path changing unit 130 as necessary, and is guided to the light irradiation unit 140. In this embodiment, the incident light adjusting section 120 may be provided after the optical path changing section 130.

The incident light adjusting section 120 includes, for example, a light amount adjusting element 122 and a polarizing element 124. The light amount adjustment element 122 receives the light 112 from the light source 110 and adjusts the intensity of the irradiation light 148 that is irradiated onto the sample 154 to be analyzed. The light amount adjustment element 122 is, for example, a neutral density (ND) filter.

Specifically, various optical filters, such as metal thin film filters and dielectric filters, can be used as the light amount adjusting element 122. By using, for example, an ND filter with a transmittance of 0.1% to 50% as the light amount adjustment element 122, the light intensity, which is the amount of incident light, can be easily adjusted.

The transmittance of the light amount adjustment element 122 may be adjusted in accordance with the intensity of scattered light of the sample to be analyzed, the detection sensitivity and exposure time of the photodetector 202 of the scattered light detection section 200, and the degree of amplification of the detection signal. Here, the amplification degree refers to an amplification factor, gain, or gain. The photodetector 202 allows the scattered light from the sample 154 to be analyzed to have a good S/N ratio, that is, the ratio of the signal signal to the noise signal, with respect to the amount of the irradiation light 148 emitted from the light irradiation section 140. It is preferable that the light amount of the incident light 126 is set so that

The polarizing element 124 included in the incident light adjustment unit 120 has a function of adjusting the polarization state of the light 112 from the light source 110 and setting the polarization state of the incident light 126, and is an optical element that adjusts the polarization state. It is configured. As the polarizing element 124, if the light 112 from the light source 110 is non-polarized light, various polarizing plates or polarizing filters can be used to set the polarization of the incident light 126.

Further, if the light 112 from the light source 110 is linearly polarized light, a 1/2 wavelength plate or the like may be used as the polarizer. By using this, the polarization direction of the incident light 126 can be set to any azimuth angle, and TM polarization (p) polarization whose electric field vector is horizontal to the optical system or TE (s) polarization whose electric field vector is perpendicular to the optical system can be set. can get. Furthermore, if the light 112 from the light source 110 is linearly polarized light, the polarization of the light 112 from the light source 110 is converted into circularly polarized light or elliptically polarized light using a quarter-wave plate or the like, and the light is irradiated. It is also possible to use it as the incident light 126 of the section 140 if necessary.

Additionally, if the intensity of the forward or backward small-angle scattered light 160 from the sample 154 to be analyzed is strongly dependent on the polarization of the incident light 126, the polarizing element 124 may be used to increase the scattered intensity of the incident light 126. It is desirable to adjust the polarization state of the incident light 126 by.

(3) Description of Optical Path Changing Unit 130 In order to actually manufacture the analyzer 100, it may be preferable to change the optical axis of the optical path from the light source 110 to the light irradiation unit 140. In such a case, it is possible to use the optical path changing unit 130. Note that the configuration shown in FIG. 1 in which the light 112 from the light source 110 is adjusted as described above in the incident light adjustment unit 120 and then the optical path of the incident light 126 is changed is an example. The optical path can be changed by providing the optical path changing unit 130 at a location where the optical path needs to be changed.

The optical path changing unit 130 is composed of an optical element such as a group of mirrors, a lens, a prism, etc., for example. For example, the optical path changing unit 130 may include a plane mirror 132 disposed at an incident angle of 45 degrees with respect to the optical axis of the light 112 from the light source 110, or an incident angle of 45 degrees with respect to the reflected light from the plane mirror 132. A plane mirror 134 is provided which is arranged in a state that the mirror 134 is parallel to the mirror 134. With this configuration, the incident light 126 with the traveling direction of the light 112 reversed can be incident on the light irradiation section 140. Standards for arranging the optical axis of the irradiated light 148 from the light irradiation section 140, the optical axis of the parallel scattered light conversion section 170, and the photodetector 202 of the scattered light detection section 200 on the optical axis 146 of the incident light 126. The axis of is set.

This makes it easy to adjust the arrangement of the sample section 150, the parallel scattered light conversion section 170, and the scattered light detection section 200. Furthermore, it is useful for reducing the size of the analyzer 100. In addition, it becomes easy to adjust the sample position adjustment mechanism 158 of the sample section 150 so that the focal position of the irradiation light 148 generated on the optical axis 146 of the incident light 126 matches the position desired to be analyzed on the sample 154. This also leads to improved adjustment accuracy. Using the optical path changing unit 130 to set the optical path to an optimal state is useful not only for downsizing the analyzer 100 but also for improving analysis accuracy.

It may be effective to irradiate the sample 154 with the irradiation light 148 of the light irradiation unit 140 having different wavelengths and analyze the difference in the small-angle scattered light 160 due to the difference in the wavelength of the irradiation light 148. In this case, an effective configuration is to set the optical axis of the incident light 126 to the sample 154 constant and to arrange a plurality of laser light sources with different wavelengths. By providing a half mirror, a total reflection mirror, a prism, etc. for each laser light source in the optical path changing unit 130, a configuration in which different wavelengths are incident on the sample 154 described above is possible.

(4) Description of the optical aperture 138 As shown in FIG. 1, the optical aperture 138 is disposed upstream of the incident light adjusting section 140 along the optical axis 146 of the incident light 126. The optical aperture 138 has a function of preventing stray light other than the irradiation light 148 from the light irradiation unit 140 from entering the sample 154 . Furthermore, it also has a function of preventing reflected light from the sample section 150, the parallel scattered light conversion section 170, etc. from entering the sample 154 again. In this embodiment, the optical diaphragm 138 includes two diaphragms 139 arranged in parallel. Each aperture of the diaphragm 139 has a slit with a larger cross-sectional shape than the cross-sectional shape of the incident light 126 for passing the incident light 126. In this embodiment, since the incident light 126 has a circular cross-sectional shape, the slit has a circular shape. The diameter of the opening, which is a slit, formed in each diaphragm 139 is larger than the diameter of the incident light 126, and has an appropriate diameter to prevent the above-mentioned stray light and reflected light. If the diameter of the light beam of the incident light 126 is, for example, 1 mmφ, the diameter of the aperture 139 is preferably 1.2 mm or more and 2 mm or less.

Even if the optical diaphragm 138 is composed of one diaphragm 139, it has the effect of preventing the above-mentioned stray light and reflected light. However, a configuration with a plurality of optical diaphragms that are perpendicular to the optical axis 146 of the light 148 irradiating the sample 154 and shifted parallel to each other at intervals provides a greater effect. In particular, as the optical diaphragm 138, a configuration in which two diaphragms having the same diameter are spaced apart is preferred. As the diaphragm 139, for example, for the incident light 126 with a diameter of 1 mmφ, two diaphragms having an aperture of approximately 1.2 mm or more and 2 mm or less in diameter are arranged at a distance of approximately 5 cm or more and 30 cm or less in the optical axis direction of the incident light. A configuration in which they are arranged in parallel has a very large effect.

(5) Description of the light irradiation section 140 The operations of the light irradiation section 140 and the sample section 150 will be explained using FIGS. 2 and 3 in addition to FIG. 1. The light irradiation unit 140 converts the incident light 126 into a light beam that is focused within the sample 154 and irradiates the sample 154 of the sample unit 150 as irradiation light 148 . Specifically, the light irradiation unit 140 includes an optical element for condensing the incident light 126 adjusted by the incident light adjustment unit 120, and a spot size of the irradiation light 148 to be irradiated onto the sample 154 and the irradiation inside the sample 154. It has a lens adjustment mechanism 144 that adjusts the focal position of the light 148, etc.

Depending on the light beam size of the incident light 126 that enters the light irradiation unit 140, the type and size of the objective lens, etc., which is a condensing lens of the light irradiation unit 140, and the shape and size of the sample cell 152 are determined. Further, depending on the light beam size of the incident light 126, the types and sizes of lenses constituting the parallel scattered light converting section 170, which will be described later, or the types and sizes of lenses constituting the scattered light condensing section 190, scattered light detection When the photodetection camera 204 is used as the photodetector 202 of the section 200 or the scattered light detection section 200, the shape and size of the light receiving section of the photodetection camera 204 are determined. Note that the scattered light detection unit 200 has a function of detecting an image and converting it into an electrical signal, and may include a light detection camera 204. A photodetection camera 204 can be used in place of the photodetector 202.

As already mentioned above, the sample section 150 includes a sample cell 152 that stores a sample 154, and a sample position adjustment mechanism 158 for moving the sample cell 152 in the X-axis, Y-axis, and Z-axis directions. The sample cell 152 is adjusted by the sample position adjustment mechanism 158 so that the minimum beam spot 157, which is the irradiation position of the irradiation light 148, matches the analysis microregion 156, which is the part to be analyzed in the sample 154 housed in the sample cell 152. , can be minutely moved along the three-dimensional axes: the X, Y, and Z axes. As a result, the irradiation light 148 is formed and irradiated so that the minimum beam spot 157 overlaps the analysis microregion 156 at the target position in the sample 154, and small-angle scattered light 160 suitable for detection is generated from the analysis microregion 156. I can do it. As described above, the sample position adjustment mechanism 158 allows the position of the sample cell 152 to be finely moved relative to the minimum beam spot 157 along the three-dimensional axis. However, it is not necessary to limit the adjustment along three-dimensional axes; for example, it is also effective to configure the sample cell 152 so that it can be moved only in the direction along the optical axis 146. Further, the position may be finely adjusted in a plane perpendicular to the optical axis 146.

The relationship between the minimum beam spot 157 of the irradiated light 148 from the light irradiation unit 140 on the sample 154 and the small-angle scattered light 160 will be explained using FIG. 3. The maximum incident angle of the irradiated light 148 entering the sample 154 is the maximum incidence angle θi, the maximum transmission angle of the transmitted light 164 passing through the sample 154 is the maximum transmission angle θ0, and the maximum small-angle scattering emitted from the sample 154 and measured Let the radiation angle of the light 167 be radiation angle θ0S. Note that each of these angles is expressed as an angle from the optical axis 146, which is the central axis of the irradiation light 148 to the sample 154.

As shown in FIG. 3, while passing through the sample 154, the irradiated light 148 incident on the sample 154 of the sample section 150 is scattered by scatterers such as particles, aggregates, and crystal grains to be measured. The light is transmitted and transmitted light 164 is emitted from the sample 154. Since the irradiation light 148 to the sample 154 is the irradiation light 148 that is incident at various incident angles within the range of a predetermined incident angle θi, the transmitted light 164 from the sample 154 is incident at various incident angles within the range of a certain emission angle θ0. radiated at an angle.

Here, the region of transmitted light 164 of sample 154 (θ<θ0) includes small-angle scattered light 160 from sample 154 in addition to transmitted light 164. Scattered light from scatterers to be measured, such as samples containing particles or aggregates, or solid thin films, is generally much weaker than transmitted light, so in the region of transmitted light 164 (θ < θ0), small-angle scattered light is It is substantially difficult to measure this light separately from the transmitted light 164. Therefore, the measurable small-angle scattered light 160 is scattered light that appears outside the region of the transmitted light 164, that is, when θ>θ0. Therefore, as the angle of the small-angle scattered light of the sample 154, in addition to the angle θ0S from the optical axis, the measured scattering angle θS measured from the maximum transmission angle θ0 is expressed by the following equation.
[Number 1]
θS=θ0S−θ0 (Formula 1)

The condensing lens 142 of the light irradiation unit 140 that forms the irradiation light 148 to be irradiated onto the sample 154 may include various types of lenses, such as a spherical plano-convex lens, a spherical biconvex lens (hereinafter simply referred to as a "spherical convex lens"), an aspherical lens, and a cylindrical surface. It is possible to use single lenses such as plano-convex lenses and cylindrical biconvex lenses (hereinafter simply referred to as "cylindrical lenses"), as well as various achromatic lenses made by bonding two or more single lenses with different optical properties. It is. The light irradiation section 140 is formed of the above-mentioned single lens, achromatic lens, or various types thereof, so as to form a minute spot-like irradiation light onto the sample 154 of the sample section 150 and efficiently irradiate the irradiation light 148. It consists of an optical system that combines lenses.

When the light from the light source 110 is a parallel (collimated) beam such as a laser beam, for example, the light beam size is formed by an optical diaphragm or the like, and the light beam is made to enter the light irradiation section 140, so that the irradiation is changed to a minute size. Light 148 is incident on sample 154. Here, the minimum beam spot 157 of the irradiation light 148 obtained by condensing, for example, a circular parallel beam of light using the above-mentioned single lens, achromatic lens, or a combination of these various lenses is used. The size W is expressed by the following formula. Note that the focal length of the condensing lens 142 is f. Further, the size W of the minimum beam spot 157 is the diameter.
[Number 2]
W=2λ/(π・NA) (Formula 2)

Here, the size W is the diameter of the minimum beam spot 157 of a circular beam within 1/e2 (=0.135) of the central maximum value of a laser beam with a Gaussian intensity distribution, and λ is the wavelength of the light. , NA (numerical aperture) is the numerical aperture of the condenser lens system. Further, the numerical aperture NA of the condensing lens 142 of the light irradiation unit 140 is NA=D0/(2f). Here, D0 is the diameter of the light beam incident on the condenser lens system (range within 1/e2 of the central maximum value). Therefore, the size W of the minimum beam spot 157 obtained with the focal length f of the condenser lens 142 is expressed by the following equation.
[Number 3]
W=4fλ/(π・D0) (Formula 3)
For example, if the condensing lens 142 of the light irradiation unit 140 is configured with a condensing lens with a focal length f = 10 mm, the incident light 126 entering the condensing lens 142 has a diameter D0 = 0.7 mm and a parallel wavelength λ = 473 nm. When the laser light beam is focused, a minimum beam spot with a diameter W of the minimum beam spot 157 of 8.6 μm is obtained.

Here, for example, when the light from the light source 110 is a laser beam, the incident light 126 that enters the condenser lens 142 becomes larger than the beam diameter at the exit aperture of the light source 110 due to the spread angle of the laser beam itself. That is, the diameter D0 of the incident light 126 that enters the condenser lens 142 is expressed by the following formula.

[Number 4]
D0=Dout+ωL (Formula 4)
Here, Dout is the beam diameter at the exit of the light source 110, ω is the spread angle (full angle) of the laser beam, and L is the distance from the exit of the light source 110 to the incident side of the light irradiation section 140. Note that the light beam diameters of D0 and Dout are within 1/e2 (=0.135) of the central maximum value of the laser light having a Gaussian intensity distribution.

For example, the value of the diameter D0 of the incident light 126 that enters the light irradiation unit 140 is Dout = 0.7 mm from a laser light source of ω = 1.2 mrad, and for example, when the distance L from the laser light source to the condenser lens 142 is 665 mm. In one case, D0=1.5 mm when calculated from the above formula.

Therefore, for example, using the condensing lens 142 with a focal length f=10 mm, a laser beam with a diameter Dout=0.7 mm outputted from the light source 110 with λ=473 nm and ω=1.2 mrad is condensed. When the distance L to the optical lens 142 is 665 mm, the diameter W of the minimum beam spot 157 obtained by condensing the light with the condenser lens 142 is W = 4.0 μm, which is smaller than the above value (8.6 μm). Thus, due to the spread of the laser light beam, the irradiation light 148 that forms the minimum beam spot 157 with a smaller size is obtained.

Here, as described above, the size W of the minimum beam spot 157 of the irradiation light 148 obtained by condensing laser light etc. with the condensing lens 142 of the light irradiation unit 140 is such that the medium around the condensing spot is air ( This is a calculated value when the refractive index n=1.00). The size of the minimum beam spot 157 in the sample 154 of the light irradiation unit 140 is because the medium has a larger refractive index than air (for example, n = 1.33 in the case of water), so the actual value of the size W is , are slightly larger than these calculated values. Therefore, as shown in FIG. 3, the size of the minimum beam spot 157 of the irradiation light 148 irradiated onto the sample 154 from the condenser lens 142 of the light irradiation unit 140 is slightly larger than the above calculated value.

The light irradiation section 140 includes a lens adjustment mechanism 144 that finely adjusts the distance between the condenser lens 142 and the sample 154 in order to irradiate the sample 154 of the sample section 150 with the irradiation light 148 . That is, the light irradiation unit 140 includes a mechanism for finely adjusting the focal position (distance) of the condensing lens 142 to the sample 154. For fine adjustment of the focal position (distance) of the condenser lens 142, for example, an optical stage equipped with a micrometer that moves the condenser lens 142 by a micrometer can be used. More specifically, for example, a precision optical stage with a moving distance of ±3 mm on which the stage mounted with the condensing lens 142 and a minimum scale of a micrometer of 10 μm can be used and is appropriate.

As shown in the examples in FIGS. 1 to 3, the maximum incident angle θi when the sample 154 is irradiated with the irradiation light 148 formed by the condenser lens 142 of the light irradiation unit 140 is The diameter D0 of the circular light beam of the incident light 126 and the focal length f of the condensing lens 142 are approximately determined as follows.
[Number 5]
θi=tan-1(D0/2f) (Formula 5)

For example, when condensing a laser beam with D0=1.5 mm using a condensing lens 142 with a focal length f=10 mm and irradiating it onto the sample 154 of the sample section 150, the maximum incident angle θi shown in FIG. is 4.3 degrees, and the sample 154 is irradiated with the irradiation light 148 with an incident angle θ<4.3 degrees.

Further, for example, when condensing a laser beam of D0=1.5 mm using the condensing lens 142 of focal length f=4 mm and irradiating it onto the sample 154 of the sample section 150, the maximum incident angle θi is 10 .8 degrees, and the measurement sample sample 154 is irradiated with the irradiation light 148 with an incident angle θ<10.8 degrees. As the focal length f of the condenser lens 142 becomes shorter, the maximum angle of incidence θi becomes larger, and the maximum angle θ0 of the transmitted light 164 also becomes larger. As a result, the measured scattering angle θS decreases. On the other hand, as described above, the size W of the minimum beam spot 157 can be narrowed, and the analysis micro region 156 can be narrowed.

The maximum incident angle θi of the irradiation light 148 irradiated onto the sample 154 by the light irradiation unit 140 is preferably in the range of 0.5 degrees to 20 degrees, and particularly preferably in the range of 0.5 degrees to 10 degrees. It is desirable to set the maximum incident angle θi of the irradiation light 148 under the above-mentioned conditions, and to cause the irradiation light 148 at an incident angle less than or equal to the maximum incidence angle θi to be incident on the sample 154 to generate scattered light from the sample 154.

More precisely, the maximum incident angle θi of the irradiation light 148 irradiated onto the sample 154 is determined by the focal length f of the condenser lens 142 of the light irradiation unit 140 and the size of the light beam of the incident light 126 that is input to the condenser lens 142. (diameter), the distance between the condensing lens 142 and the sample 154, the refractive index of the sample 154 itself, the length in the optical axis direction of the transmitted light passing through the sample 154 (optical path length), and the sample that accommodates the sample 154. It is determined by the shape of the cell 152, the material (refractive index) of its constituent members, etc.

The maximum angle of incidence θi shown in FIG. 3 may be smaller than 0.5°. θi=0°, that is, the light incident on the sample 154 of the measurement sample section 4 may be parallel light (collimated light). In this case, the size of the light beam irradiated onto the sample 154 is adjusted using a group of optical elements for beam light formation, which is a combination of optical elements such as a lens, a pinhole, and an optical diaphragm, which constitute the light irradiation section 140. Then, parallel light (collimated light) is made to enter the sample 154.

As a specific condensing lens 142, an objective lens having a short focal length and high NA is particularly suitable. More specifically, an objective lens with a relatively long working distance WD and an infinity correction is particularly preferable. As the condensing lens 142, it is also possible to use a finite type microscope objective lens that can form an enlarged image even in the absence of an imaging lens.

For example, the NA values of these objective lenses suitable as the condenser lens 142 are 0.13 for a 5x objective lens, 0.3 for a 10x objective lens, and 0 for a 20x objective lens, respectively. .4, 0.55 for a 50x objective, and 0.8 for a 100x objective. Further, for example, the working distance WD of these objective lenses as the suitable condensing lens 142 is comparatively about 2 mm (in the case of a 100x objective lens) to 11.6 mm (in the case of a 5x objective lens). long.

As the condensing lens 142 of the light irradiation unit 140, for example, a long working distance objective lens with infinity correction, a 20x objective lens (focal length f is 10 mm, working distance WD is 11.1 mm, NA is 0.4, A 50x objective lens (with f of 4 mm, WD of 8.2 mm, NA of 0.55, and resolution of 0.5 μm) is suitable.

(6) Description of sample section 150 The sample section 150 will be explained. The sample section 150 includes a sample cell 152 that stores a sample 154, and a sample position adjustment mechanism 158 that supports the sample cell 152 and finely moves and adjusts the position of the sample cell 152 with respect to the optical axis 146 of the irradiation light 148. have The sample position adjustment mechanism 158 moves the analysis micro region 156, which irradiates the sample 154 with the irradiation light 148 formed by the condensing lens 142 of the light irradiation unit 140, on a two-dimensional plane perpendicular to the optical axis 146 or in a three-dimensional direction. It has the ability to move minutely. Furthermore, although not shown, it has a function of controlling the temperature of the sample 154.

When the sample 154 is a liquid, the sample cell 152 is a container that accommodates the liquid and can transmit scattered light from the sample 154. When the sample 154 is a solid such as a thin film, the sample 154 may have a structure that accommodates and fixes the solid and allows scattered light from the sample 154 to pass through. Furthermore, for example, when the measurement target is a thin film formed on a transparent substrate such as a glass substrate, the substrate itself on which the thin film is formed functions as the sample cell 152, and the thin film formed on the substrate etc. serve as the sample 154.

The installation position of the sample cell 152 can be adjusted by a sample position adjustment mechanism 158. After the adjustment is completed, the measurement operation begins. It is desirable to find a measurement spot that is desirable for analysis in the sample 154, and at the desired measurement spot, measure data that can be used for both DLS measurement and SLS measurement using the scattered light detection unit 200 and import it into the storage device 290. Therefore, as will be explained below, the light irradiation section 140 irradiates the irradiation light 148 for adjustment, and the scattered light detection section 200 measures the small-angle scattered light 160 generated by the irradiation of the irradiation light 148. The based image is displayed as an adjustment image on the input/output device 266 of the analysis unit 250. While viewing this adjustment image, it is possible to move the sample cell 152, for example, on a plane perpendicular to the optical axis 146, and find the irradiation position where the image displayed on the adjustment image becomes a desired image. . With this adjustment image, adjustment can be performed very efficiently and optimally set as the analysis microregion 156.

When the sample 154 is a liquid, the sample cell 152 is preferably a rectangular cell in which the entrance surface 151 and the transmission surface 153 of the sample cell 152 shown in FIG. 3 are flat. In addition, in order to irradiate the liquid sample 154 with the irradiation light 148 from the light irradiation unit 140 in a spot shape and efficiently generate and measure the small-angle scattered light 160, it is necessary to adjust the direction of the optical axis of the irradiation light 148 passing through the sample 154. It is preferable that the optical path length LS of the sample 154 is as short as possible. As the optical path length LS of the sample 154 becomes shorter, the spot beam of the irradiation light 148 formed by the condensing lens 142 of the light irradiation section 140 hardly spreads inside the sample 154, and the scattered light is scattered from a local part of the sample 154. The sample 154 can be released for localized analysis within the sample 154.

Therefore, it is preferable that the sample cell 152 that accommodates the sample 154 has a short optical path length LS, which is the length in the optical axis direction, that is, the sample cell 152 is as thin as possible. For example, the optical path length LS of the sample 154 is preferably 1 mm or less, and more preferably 0.5 mm or less. Therefore, the length between the entrance surface 151 and the transmission surface 153 of the sample cell 152 is preferably about the same size, that is, 1 mm or less, and more preferably 0.5 mm or less. Further, the volume of the sample 154 used is preferably from several hundred μl or less to several tens of μl or less. The above l represents liter.

Therefore, as a sample cell 152 containing a liquid sample 154, for example, a flat spacer of a specific thickness is sandwiched between two transparent flat substrates, and the sample 154 is injected between the substrates formed by the spacer. Preferably, a sample cell 152 of sandwich type structure is used. In the case of forward small-angle scattered light measurement, the irradiated light 148 from the light irradiation section 140 enters from one of the two transparent flat substrates (incident side), passes through the sample 154, and then passes through the sample 154. Scattered light emitted from one substrate side is measured. In addition, in the case of small-angle backward scattered light measurement, the irradiation light 148 from the light irradiation section 140 enters from one of the two transparent flat substrates, and the scattered light is emitted from this same substrate side. Measure. That is, on one substrate side of the sample 154, light is incident on the sample 154 and scattered light is emitted.

Specifically, it is preferable to use an optical material with excellent transparency in the wavelength range of the light to be measured on the incident light side and the scattered light emission side as the above-mentioned transparent flat substrate. For this reason, various transparent glasses, transparent plastics, etc. are suitable.

When the sample 154 is a solution containing biomolecules such as proteins, peptides, and nucleic acids or their aggregates, the scattered light is significantly large in a small scattering angle range of 10 degrees or less, especially in a scattering angle of 8 degrees or less. Scattered light is noticeable. Since the scattered light from the sample 154 passes through the transparent sample cell 152 and is emitted to the outside, it is affected by refraction at the incident surface 151 and the transmitting surface 153.

Further, as described above, the condensing lens 142 of the light irradiation unit 140 allows the maximum incident angle θi to preferably be in the range of 0.5 degrees to 20 degrees, particularly the maximum incident angle θi in the range of 0.5 degrees to 10 degrees. Preferably, the illumination light 148 is incident on the sample 154 at an angle. The incident angle on the incident light side of the sample 154 is also preferably a small angle, similar to the scattering angle of the small-angle scattered light 160. In this embodiment, since the incident surface 151 and the transmitting surface 153 of the sample cell 152 are flat, if the thickness thereof is thin to some extent, the incident angle is 20 degrees or less, especially 10 degrees or less, which is a small angle incidence. Therefore, when the scattering angle is small, that is, 10 degrees or less, there is no particular need for correction in the refraction angle due to refraction as described above. In other words, if the sample cell 152 is a flat plate with a certain degree of thinness, it is especially necessary to correct the scattering angle in consideration of the refraction angle in the sample cell 152 when measuring small-angle scattered light for the irradiation light 148 at a small incident angle. It will no longer be.

Therefore, in the measurement of the small-angle scattered light 160, it is preferable that the flat plates of the entrance surface 151 and the transmission surface 153 that constitute the sample cell 152 are as thin as possible. Specifically, in the case of measuring the small-angle scattered light 160 in the irradiation light 148 with small-angle incidence, the thickness tin of the flat plate of the incident surface 151 of the sample cell 152 and the thickness tout of the flat plate of the transmitting surface 153 are The diameter D0 of the incident light 126 entering the condenser lens 142 is preferably within the range of 1/10 to 1/5. That is, the conditions of thickness tin<D0/10 to D0/5 and thickness tout<D0/10 to D0/5 are preferable. In particular, it is more preferable that the thickness tin<D0/10 and the thickness tout<D0/10.

In addition, the occurrence of forward small-angle scattered light with a small scattering angle is remarkable in solutions containing biomolecules such as proteins, peptides, and nucleic acids, or their aggregates, and in solid thin films that form crystal grains such as polymers such as plastics. Depending on the structure and optical arrangement of the sample 154, scattered light may be emitted on the same side as the incident side of the irradiation light 148. In this case, scattered light from the sample 154 is detected on the incident side of the irradiation light 148. That is, depending on the sample 154, the small-angle backward scattered light may appear more prominently than the small-angle forward scattered light. Small-angle backward scattered light is a scattering angle in which the transmitted light 164 of the irradiated light 148 from the optical axis 146 ranges from 170 degrees or more to 180 degrees (inverted direction of incident light), particularly scattering from 172 degrees to 180 degrees. This is the scattered light at the corner.

Specifically, the small-angle backward scattered light is, for example, when the light 148 irradiated onto the sample 154 is reflected by the wall on the opposite side of the sample cell 152 housing the sample 154, and the reflected light is reflected by the sample 154 as small-angle scattered light. is generated, and scattered light is emitted on the same side as the incident side of the irradiation light 148. In this case, the backward small-angle scattered light will be measured. For example, the flat plate on the incident side of the sample cell 152, which is composed of two transparent flat plate-shaped substrates, also serves as the scattered light output side, so that the forward small-angle scattered light is measured. Similar to the measurement of , thinner is preferable. However, the other flat plate on the opposite side may be thicker as long as it can efficiently reflect the transmitted light 164 passing through the measurement sample. In the sample cell 152 used for measuring small-angle backward scattered light, the flat plate provided on the front side of the irradiated light 148 may be made of any material that reflects light, such as glass or various plastics. The front flat plate used needs to have a smooth surface so as not to emit scattered light from itself, and preferably reflects the transmitted light well.

Further, the flat plates of the entrance surface 151 and the transmission surface 153 that constitute the sample cell 152 may be made of transparent members of the same material and the same thickness, or may be made of different materials and of different thicknesses. When the flat substrates on the incident side and the scattered light output side are constructed of transparent members of the same material and thickness, there is no distinction between the incident surface 151 and the transmitting surface 153 in the sample cell 152. That is, the irradiation light 148 may be incident from either side. In this case, as the flat plate of the sample cell 152, it is possible to use, for example, a cover glass of the same standard with a thickness of 0.2 mm or less.

As the flat spacer of the sample cell 152, glass, polystyrene, polypropylene, fluororubber, silicone rubber, etc. are suitable. Silicone rubber is particularly preferred, and sheets or films are used. Silicone rubber has good adhesion to glass substrates, etc., and when analyzing the sample cell 152 by batch method, within a few hours, the concentration of various solutes in the sample 154 will hardly change due to solvent evaporation from the gap. It's not a problem. However, when using the measurement sample solution while being held in the sample cell 152 for a long period of time (several tens of hours or more), silicone grease or fluoride is added between the spacer and the flat substrate to further prevent liquid leakage. It is preferable to seal with grease or the like.

When the solvent of the sample 154 is an organic solvent such as ethanol or acetonitrile, the silicone rubber used as the spacer of the sample cell 152 has weak adhesion to the glass substrate or the like. In that case, seal the gap between the spacer and the flat substrate with silicone grease, fluorine grease, etc., or use a clip or the like to hold the flat plate of the sample cell 152 under a certain degree of pressure. Improves the adhesion between rubber and glass substrates, etc., and prevents solvent evaporation and liquid leakage from gaps. Alternatively, the sample position adjustment mechanism 158 that holds the sample cell 152 applies a certain amount of pressure to the flat substrate to hold it.

As described above, the spacer and two flat plates that make up the sample 154 function as the sample cell 152 that holds the sample 154. For this purpose, for example, a groove of an arbitrary shape such as a circle, an oval, or a rectangle is formed in a flat spacer, and the sample 154 is placed in the groove space formed by sandwiching this spacer between two flat plates. It can be stored. That is, in the sample cell 152, a sample 154 is injected into the groove space and held therein.

Analysis of a sample 154, such as a solution or a thin solid film, is often performed as a homogeneous and uniform sample. Therefore, analysis of the sample 154 by measuring the small-angle scattered light 160 provides information about the movement and structure of particles, aggregates, etc. in the sample 154 on a part or all of the sample irradiated with light. Information can be obtained. However, the following analysis of the sample 154 requires not only average information as described above, but also local analysis. In other words, for example, in samples where the concentration of constituent components locally differs in the liquid due to liquid-liquid phase separation, or in crystallization solutions such as proteins, peptides, and nucleic acids, the formation of aggregates or crystal nuclei may occur in the solution. Samples in which crystal grains are formed unevenly in a thin film such as a polymer, etc. are to be analyzed. When such a sample 154 is to be analyzed, the distribution within the measurement sample on a microscopic scale is measured by measuring the scattered light.

The thickness tsp of the spacer used in the sample section 150 corresponds to the optical path length in the sample 154. The irradiated light 148 from the condenser lens 142 is allowed to pass through the sample 154 while maintaining an appropriate minute spot without being expanded onto the sample 154 as much as possible, and the scattered light from the minute irradiated portion of the sample 154 is efficiently radiated. Therefore, it is preferable that the optical path length of the sample 154 to be measured is short, and that the capacity of the sample 154 is also very small. Specifically, it is preferable that the volume of the sample 154 to be measured is small, for example, 25 μl or less. In this case, the thickness tsp of the spacer is preferably about the diameter of the incident light 126 that enters the condenser lens 142 of the light irradiation section 140, for example, 1 mm or less, particularly 0.5 mm or less.

In addition to scattering light measurement with the sample 154 sealed inside the sample section 150, that is, sample analysis in a batch method, sample liquid, aggregating agent, or crystallizing agent is introduced into the sample cell 152 from the outside during analysis. It is also possible to configure the device so that solutions such as these can be injected and discharged. In this case, the sample cell 152 may be provided with an insertion port, a discharge port, etc. for injecting or discharging the liquid. In that case, during the analysis of the sample 154 by measuring the scattered light, it is possible to perform the analysis by adjusting or changing the concentration of each substance.

It is also conceivable to use a commercially available rectangular or cylindrical solution cell for light absorption measurement as the sample cell 152. Many commercially available solution cells have a long optical path length LS of, for example, 5 mm to 1 cm, and require several ml or more of sample liquid, making it difficult to apply them as the sample cell 152. However, if the optical path length of a commercially available solution cell is particularly short, for example about 1 mm or less, it is also possible to use it. However, in this case, in the measurement of the forward small-angle scattered light or the backward small-angle scattered light, the refraction in the sample cell 152 is It is necessary to correct the incident angle and scattering angle of the condensed light, taking into account the angle.

The sample position adjustment mechanism 158 that supports the sample cell 152 includes a rotation mechanism and a tilting mechanism for adjusting the angle of incidence on the sample 154, a mechanism for moving the position relative to the optical axis that is the central axis of the incident light beam, and the like.

The angle of incidence of the center of the incident light on the sample cell 152 is perpendicular to the central axis of the irradiated light 148 by the condenser lens 142 of the light irradiation unit 140, that is, the optical axis 146 of the incident light 126 (incident angle 0°). is particularly preferred. Further, it is sufficient that the angle of incidence is small, for example, 1 to 2 degrees or less near the angle of incidence of normal incidence, and the reflected light of the sample cell 152 becomes stray light and is received by the photodetector 202 of the scattered light detection unit 200. The angle of incidence is adjusted by the sample position adjustment mechanism 158 so that the sample does not enter the area. To adjust the angle of incidence of the condenser lens 142, a rotating stage, a tilting stage, or the like can be used. Further, the structure is such that the incident angle can be set to, for example, 1 to 2 degrees or less when fixing the sample cell 152 to the sample position adjustment mechanism 158 without using a rotation stage or a tilting stage. All you have to do is stay there.

The sample position adjustment mechanism 158 that supports the sample cell 152 has a structure that allows the sample cell 152 to be moved two-dimensionally in a direction perpendicular to the optical axis 146 of the irradiation light 148, so that a minute spot of incident light is generated in the in-plane direction of the sample 154. The beam is relatively scanned to measure scattered light from a minute area of the sample 154. This allows analysis of the sample 154 on a microscopic scale.

Specifically, the micro-movement of the sample cell 152 is performed using, for example, a two-axis stage with an X-axis and a Y-axis, with a support holder placed on a two-axis stage with an X-axis and an XY-axis. Furthermore, the support holder is installed on the Z-axis stage in the optical axis direction, that is, the support holder is installed on the three-axis stage of the X-axis, Y-axis, and Z-axis, and the sample cell 152 is minutely moved. Good too. In addition, when investigating a certain one-dimensional distribution in local analysis of the sample 154, for example, a support holder is installed on an X-axis stage or a Y-axis stage, and the sample cell 154 is may be moved slightly.

Further, as the support holder, an XY two-axis stage or an XYZ three-axis stage for minutely moving the sample cell 152 may be used. In this case, the sample cell 152 is placed and held on an XY two-axis stage or an XYZ three-axis stage.

As described above, the sample position adjustment mechanism 158, which is a mechanism for minutely moving the sample cell 152, may be, for example, a two-axis X-axis and Y-axis stage with a through hole through which light can pass, or an X-axis and Y-axis stage. A 3-axis stage for axis and Z-axis can be used. A manual stage that moves linearly using a micrometer as a multi-axis stage with two axes (X and Y axes) or three axes (X, Y, and Z axes), for example, a table surface size of 120 mm x 120 mm. A multi-axis stage with an amount of ±20 mm and a minimum micrometer scale of 10 μm can be used. If the minimum beam spot 157 of the irradiation light 148 irradiated onto the sample 154 by the condensing lens 142 of the light irradiation unit 140 is, for example, 10 μm or less in diameter, by moving the stage with a micrometer, for example, by a minimum of 10 μm, The irradiation position within the sample 154 can be scanned. Therefore, the scattered light measurement enables local analysis of the sample 154 on a microscopic scale.

Further, when performing microscopic analysis with a resolution of about 10 μm or less by scanning the sample 154 with ultra-fine movement, for example, an ultra-precision stage driven by a piezo element can be used as the sample position adjustment mechanism 158. As an ultra-precision stage driven by a piezo element, for example, a table surface size of 120 mm x 120 mm, a movement amount of 100 μm, and a minimum positioning resolution of 0.01 μm can be used. Here, in scanning the sample 154, a multi-axis stage may be configured and used by combining a stage with a micrometer and a stage driven by a piezo element. In this case, for example, when scanning the sample 154, a stage with a micrometer is used for fine movement of 10 μm or more, and a piezo element driven stage is used for fine movement of about 10 μm or less.

The objects of analysis, such as small and medium molecules such as peptides and nucleic acids, biopolymers such as proteins, particles and aggregates of inorganic substances such as metals and plastics, are analyzed by the thermal movement of solvent molecules in liquids, such as water molecules in aqueous solutions. In response to collisions, it undergoes Brownian motion, which is random diffusion. Such Brownian motion is sensitive to the temperature T of the liquid in which particles and aggregates are present. For example, the diffusion coefficient D [m2/s] of Brownian motion of spherical particles is expressed by the Stokes-Einstein equation as shown below, and is proportional to the temperature T [K] of the liquid.
[Number 6]
D=kBT/(6πηa) (6)
Here, η [kg/m・s] is the viscosity coefficient (viscosity coefficient, viscosity) of the liquid medium, a [m] is the radius of particles or aggregates, and kB is Boltzmann's constant (kB = 1.380 × 10-23 J /K).
By evaluating the diffusion coefficient D of the scatterer as a spherical particle from dynamic light scattering (DLS) measurement, the size (radius) a of the spherical particle can be calculated as a=kBT/(6πηD) using the above Stokes-Einstein formula. ) can be estimated. Here, the viscosity η of the liquid medium and the temperature T of the liquid are measured by other appropriate methods, and their measured values are used. Note that it is preferable to measure the scattered light while the temperature of the liquid sample 154 is adjusted and controlled to a predetermined constant temperature.

On the other hand, there is a need to change the temperature of the sample 154 during measurement of scattered light and analyze the state of particles and aggregates in the liquid in real time. Specifically, this is the case, for example, when analyzing the temperature-dependent process of formation of aggregates in solutions of proteins, peptides, etc. For example, while measuring the scattered light from the sample, the temperature of the sample is raised or lowered to analyze and evaluate the state of aggregate formation in real time. In such an analysis, scattered light is measured while changing the temperature of the liquid sample 154 at a predetermined heating rate or cooling rate. Therefore, the temperature of the liquid sample 154 can be changed at a predetermined heating rate or a predetermined cooling rate, and while the temperature of the sample 154 is adjusted and controlled, the scattered light generated by the sample 154 is measured. It becomes possible to do this.

In addition, in the case of a thin film that forms crystal grains of polymers such as plastics, which is another object of analysis, the formation of crystal grains changes sensitively to temperature. Therefore, similarly to the liquid sample 154, the scattered light generated by the sample 154 is measured while the temperature of the sample 154, such as a thin film, is controlled to be maintained at a predetermined constant temperature. Furthermore, in the case of a sample 154 such as a thin film, the temperature of the sample 154 is changed at a predetermined heating rate or a predetermined cooling rate, and the scattered light generated by the sample 154 under such an environment is measured. This makes it possible to analyze a sample 154 such as a thin film.

The light irradiation unit 140 includes a mechanism that adjusts and controls the temperature of the sample 154. As a device for adjusting and controlling the temperature of the sample 154, various temperature controllers, cooling/heating stages, etc. can be used. These temperature controllers change the temperature by heating with a heater, cooling with a refrigerant, heating or cooling with a Peltier element, etc., and measure the temperature of the object with a temperature sensor such as a platinum resistor or semiconductor. The temperature is adjusted and controlled using, for example, a PID control method while feeding back to the heating and cooling mechanism. Since the temperature of the sample 154 is adjusted and controlled via the sample cell 152 that houses the sample section 150, the temperature of the sample 154 is adjusted and controlled appropriately, taking into consideration the thermal properties (heat capacity, thermal conductivity, etc.) of the sample section 150 including the sample 154. Use a suitable temperature controller.

When the temperature controller cools the sample 154 below room temperature, it is necessary to prevent condensation. Water droplets adhere to the sample cell 152 due to dew condensation due to cooling, and scattered light from the water droplets becomes strong back noise, making it difficult to measure the scattered light of the sample 154. Therefore, it is preferable to introduce an inert gas such as nitrogen or argon into the outer peripheral portion of the sample cell 152, and an inert gas inlet is provided at a portion where the sample cell 152 of the temperature controller is attached. In addition, the introduction of an inert gas has the effect of preventing oxidation that may occur when heating samples such as thin films that form crystal grains of polymers such as plastics, so it is especially important when used at high temperatures. Introduction of active gas is preferred.

As a device for adjusting and controlling the temperature of the sample 154, the cooling/heating stage using a Peltier element or the like is compact because it can perform heating and cooling with a small Peltier element, and is suitable for the small sample cell 152. Particularly preferred. More specifically, for example, a Peltier cooling and heating stage for a microscope is suitable. A Peltier cooling/heating stage for a microscope, for example, heats and cools a sample using a combination of a Peltier element and a water circulation unit.

When the liquid sample 154 is the measurement target, the temperature of the sample can be adjusted and controlled with an accuracy of ±0.05°C in the temperature range of -20°C to 120°C, for example. Further, the temperature increase/decrease rate can be set at, for example, 0.01 to 20° C./min.

In addition, when a solid object such as a thin film is to be measured, for example, in the temperature range of -190°C or room temperature to 600°C, the temperature accuracy is ±0.2°C (-190°C to room temperature), ±0.05°C ( The sample temperature can be adjusted and controlled from room temperature to 600℃. Further, the temperature raising/lowering rate can be set to, for example, 0.01°C/min to 20°C/min (-190°C to room temperature) and 0.01°C/min to 150°C/min (room temperature to 600°C).

The Peltier type cooling and heating stage for a microscope is provided with an optical window that enables microscopic observation of the sample 154, and is compatible with an objective lens for microscopic observation. The Peltier cooling/heating stage for a microscope is compatible with objective lenses with long working distances of, for example, 20x, 50x, and 100x, and enables microscopic observation of the sample while controlling the sample temperature. It is suitable as a temperature controller for the sample 154 in the particle/aggregate analysis device.

A sample cell 152 that stores a liquid sample 154 is configured to adjust and control the temperature of the sample 154. The temperature of the sample 154 may be controlled by directly attaching and holding it to a cooling/heating part such as a cooling/heating stage. That is, in this case, the cooling/heating stage itself functions as a support holder for the sample section 150 and holds the sample cell 152. Further, when the sample 154 is a solid object such as a thin film, the temperature of the sample 154 may be controlled by directly attaching and holding the sample 154 to a cooling/heating part of the cooling/heating stage or the like.

The cooling/heating stage, etc., which is a device for adjusting and controlling the temperature of the sample 154, performs microscopic scanning of the sample cell 152 in order to microscopically analyze the sample 154 by measuring scattered light. It may also be mounted on a multi-axis stage with three axes: the X-axis, Y-axis, and Z-axis. Further, the cooling/heating stage or the like may be provided with a mechanism for slightly moving the sample cell 152 or the sample 154 itself, for example, in two axial directions of the X axis and the Y axis. Specifically, for example, microscopic analysis can be performed by microscanning the sample cell 152 or the measurement sample using a micrometer stage built into a cooling/heating stage or the like, and measuring scattered light from a local region of the sample 154. can.

(7) Description of the parallel scattered light converter 170 The parallel scattered light converter 170 converts the small angle scattered light 160 of the scattered light by the sample 154 emitted from the beam of the irradiation light 148 irradiated onto the sample part 150. It is captured within the range of the measurement scattering angle θS and converted into parallel small-angle scattered light 168 parallel to the optical axis 146. Although this embodiment describes the case where the forward small-angle scattered light 160 is used, the concept is the same for the backward small-angle scattered light, and similar actions and effects can be obtained. Specifically, the sample 154 is irradiated with irradiation light 148 that is focused to a predetermined size by the condensing lens 142 of the light irradiation unit 140 . In addition to the transmitted light 164 from the sample 154, small angle scattered light is emitted forward. At this time, small-angle scattered light is also emitted rearward at the same time, but as mentioned above, in this example, the configuration using small-angle scattered light emitted forward is used to measure scattered light generated forward and backward. This will be explained as a typical example of an analyzer that performs analysis. Assuming that the irradiation light 148 is irradiated onto the minimum beam spot 157 of the sample 154, the small-angle scattered light 160 emitted from the minimum beam spot 157 within the range of the measured scattering angle θS enters the lens 172, and is caused by the lens 172 to It is converted into parallel small angle scattered light 168 which is parallel light (collimated light). The diameter of the minimum beam spot 157 is, for example, in the range of several tens of μm to several hundred μm, and is preferably in the range of 10 μm or more and 100 μm or less.

The parallel scattered light converter 170 converts the parallel small angle scattered light 168 into parallel small angle scattered light 168, and the parallel small angle scattered light 168 enters the scattered light detector 200, where the intensity of the parallel small angle scattered light 168 is detected. be done. Note that when using small-angle scattered light generated at the rear, a parallel scattered light converter 170 (not shown) arranged at the rear converts it into light parallel to the optical axis 146, and its intensity is transmitted to the scattered light detector 200. Detected with similar equipment. Specifically, the scattered light detection unit 200 measures the parallel small-angle scattered light 168 at once as an intensity distribution in a predetermined scattering angle range, or continuously measures the intensity distribution in a predetermined time range. . Note that the scattered light detection unit 200 may capture a moving image of the parallel forward small-angle scattered light or the parallel backward small-angle scattered light as a parallel forward small-angle scattered light image or a parallel backward small-angle scattered light image, respectively.

The measurement results by the scattered light detection unit 200 are sent to the analysis unit 250, and the analysis unit 250 determines the state of particles and aggregates in the sample 154 from the intensity distribution of the parallel small-angle scattered light 168 measured by the scattered light detection unit 200. To analyze. For this reason, the analysis unit 250 temporarily stores the measurement data of the parallel small-angle scattered light 168 measured by the scattered light detection unit 200 in the storage device 290 of the arithmetic processing unit 270, and stores the measurement data in the storage device 290 as necessary. Read from and analyze.

The parallel scattered light converter 170 includes a lens 172 and a support device 174 that supports the lens 172. The support device 174 includes, for example, a mechanism for supporting the lens 172 and adjusting the position and tilt angle of the lens 172.

The lens 172 of the parallel scattered light conversion unit 170 may be a single lens such as various lenses, such as a spherical plano-convex lens, a spherical biconvex lens, a cylindrical plano-convex lens, or a cylindrical biconvex lens (hereinafter simply referred to as a "cylindrical lens"). , and various achromatic lenses in which two or more single lenses with different optical properties are bonded together can be used. The lens 172 is an optical system such as the above-mentioned single lens, an achromatic lens, or a combination of these various lenses, so as to efficiently take in scattered light spread over a predetermined scattering angle range and convert it into parallel light. It consists of

The lens 172 of the parallel scattered light converter 170 is arranged between the condenser lens 142 of the light irradiator 140 and the photodetector 202 of the scattered light detector 200, and converts the small angle scattered light 160 from the sample 154 into an optical axis. 146 is placed at a suitable position to convert the light into parallel small-angle scattered light 168. The scattered light detection unit 200 is arranged so that the parallel small-angle scattered light 168 converted by the lens 172 is perpendicularly incident on the detection surface of the photodetector 202 of the scattered light detection unit 200. That is, the photodetector 202 of the scattered light detection unit 200 is arranged perpendicular to the optical axis 146.

As the lens constituting the parallel scattered light conversion section 170, an achromatic lens with little spherical aberration is suitable. For example, when a circular achromatic lens is used, the scattered light emitted from the irradiation area of the small circular spot light of the sample 154, the minimum beam spot 157 shown in FIG. Parallel small-angle scattered light 168 can be obtained relatively easily. Further, as the lens 172 of the parallel scattered light converter 170, a shape other than circular, for example, an achromatic cylinder lens having a cylindrical shape can be used. When a single cylindrical lens or an achromatic cylinder lens is used, parallel small-angle scattered light 168 is obtained that is collimated only in the direction of the cylindrical curved surface.

In the configuration shown in FIG. 2, if the distance from the sample cell 152 of the sample section 150 to the lens 172 of the parallel scattered light conversion section 170 is the distance LB, the distance LB is the focal length f of the lens 172, the incident angle of the incident light, and the distance LB. It depends on the range, the beam size of the incident light, the size of the illuminating light 148 on the sample 154, the structure of the sample cell 152, etc. Among these factors, it largely depends on the focal length f of the lens 172 used in the parallel scattered light converter 170. The position where the lens 172 of the parallel scattered light converter 170 is arranged is such that the distance LB is theoretically equal to the focal length f, assuming that the small angle scattered light 160 is emitted from the analysis micro region 156 of the sample 154.

However, the most appropriate arrangement of the lens 172 of the parallel scattered light converter 170 is a position where the lens 172 causes the small angle scattered light 160 from the sample 154 to be parallel (collimated) to the optical axis 146. Specifically, the scattered light that has passed through the lens 172 of the parallel scattered light converter 170 becomes a parallel image, and is placed at a position where it becomes a parallel small-angle scattered light image. In other words, the position of the lens from which the parallel small-angle scattered light 168 is obtained is the position at which the lens 172 of the parallel scattered light converter 170 is arranged.

More specifically, as the lens 172 constituting the parallel scattered light converter 170, a circular achromatic lens having a diameter of 50 mm and a focal length f in a range of 50 mm to 150 mm can be used, for example. For circular lenses with the same diameter, the shorter the focal length, the wider the scattering angle range of scattered light can be taken in from the sample 154.

For example, for a sample 154 whose measurement target is latex particles with a diameter of 0.1 μm dispersed in pure water, a 20x infinity-corrected objective lens is used as the condenser lens 142 of the light irradiation unit 140, for example. Note that the focal length f of this objective lens is 10 mm, and the numerical aperture NA is 0.4. In FIG. 2, the distance LA from the condenser lens 142 to the sample 154 is, for example, 10 mm. Irradiation light 148 having a maximum incident angle θi of 4.3 degrees shown in FIG. 3 is incident on the sample cell 152. Here, the sample cell 152 that houses the sample 154 is constructed by sandwiching a 0.5 mm thick silicone rubber sheet with a 7 mm diameter hole between two 0.15 mm thick microscope cover glasses. are doing. In this case, as the lens 172 of the parallel scattered light converter 170, for example, a circular achromatic lens with a focal length f of 75 mm and a diameter of 50 mm is arranged at a position where a distance LB, which is slightly shorter than the focal length of 75 mm, is 68 mm. The lens 172 can take in from the sample 154 small-angle forward scattered light in an angular range in which the radiation angle θ0S is smaller than 20 degrees, and convert it into parallel small-angle forward scattered light.

In this embodiment, an objective lens with a short focal length and a large NA can be used as the lens 172 of the parallel scattered light converter 170. In particular, an objective lens with a relatively long working distance WD and an infinity correction is suitable. When an objective lens is used as the lens 172 of the parallel scattered light converter 170 to accurately convert the small angle scattered light 160 from the sample 154 of the sample section 150 into the parallel small angle scattered light 168 using the objective lens, it is difficult to accurately convert the small angle scattered light 160 from the sample 154 of the sample section 150 into the parallel small angle scattered light 168 described above. Compared to a lens, especially an achromatic lens with a long focal length f, for example, a focal length f of 75 mm, the focal length of the objective lens is short, making it difficult to adjust the position of the objective lens. However, a precision optical stage or the like, such as a multi-axis precision optical stage using a micrometer, is used for the support device 174 that has a mechanism for adjusting the position of the lens 172 of the parallel scattered light converter 170, so that the minute position of the objective lens can be adjusted. By adopting a structure that allows fine adjustment of the position, especially the position of the objective lens that determines the distance between the sample cell 152 and the objective lens used for the lens 172, the small-angle scattered light 160 from the sample 154 of the sample section 150 can be , which can be accurately converted using an objective lens. That is, accurate parallel small-angle scattered light 168 can be obtained.

Note that a configuration may be adopted in which the small-angle scattered light 160 is directly guided to the light receiving section of the photodetector 202 of the scattered light detection section 200 without using the parallel scattered light conversion section 170. That is, it is also possible to adopt a configuration in which the small-angle scattered light 160 from the light irradiation section 140 is measured by the scattered light detection section 200 at one time without using the parallel scattered light conversion section 170.

(8) Description of transmitted light shielding section 176 The transmitted light 164 that has passed through the sample 154 has an extremely high intensity compared to the scattered light from the sample 154. Therefore, in order to measure weak scattered light by the photodetector 202 of the scattered light detection section 200, it is necessary to avoid overscaling of the photodetection signal due to strong transmitted light entering the scattered light detection section 200. In addition, the transmitted light causes stray light to enter the scattered light detection section 200, which impedes measurement of weak scattered light. Therefore, as shown in FIGS. 1 and 2, it is preferable to prevent the transmitted light 164 from the sample 154 from entering the scattered light detection section 200 using a transmitted light shielding section 176. That is, the transmitted light blocking section 176 blocks the transmitted light 164 of the sample 154 from entering the scattered light detection section 200 or attenuates the intensity of the transmitted light 164.

The transmitted light 164 from the sample 154 passes through the lens 172 of the parallel scattered light converter 170, similar to the scattered light from the sample 154, especially the forward small-angle scattered light 160, and thus proceeds under the lens action of the lens 172. . That is, the irradiation light 148 formed by the condensing lens 142 of the light irradiation unit 140 enters the sample 154, and the transmitted light 164 spreads at various radiation angles within a predetermined maximum transmission angle θ0 and hits the sample. 154 and is captured by lens 172 as well as scattered light from sample 154 . Then, the transmitted light 164 is converted into parallel transmitted light 166 by the lens 172, similar to the scattered light from the sample 154, and the scattered light 164 is converted into parallel transmitted light 166, which is parallelized by the lens 172, and in parallel with the parallel small-angle scattered light 168. Proceed to the photodetector 202 of the detection unit 200. Therefore, the transmitted light shielding section 176 prevents the transmitted light 164 from the sample 154 from entering the photodetector 202 of the scattered light detection section 200. Preferably, it is mounted on axis 146. Particularly when measuring small-angle scattered light 160 from the sample 154, the transmitted light shielding section 176 is arranged, for example, between the lens 172 and the detection surface of the scattered light detection section 200, or between the sample cell 152 and the lens 172. do.

The transmitted light shielding section 176 can use various types of absorbers, mirrors, and the like. Furthermore, the transmitted light shielding section 176 needs to not generate scattered light. There are no particular restrictions on the material of the transmitted light shielding section 176 as long as it is made of a material that has a function of blocking transmitted light or attenuates the intensity of transmitted light and does not generate scattered light. For example, paper, wood, metal, plastic, glass, etc. can be used as the material for the transmitted light shielding section 176. In order to enhance the effect of blocking transmitted light or attenuating transmitted light, it is preferable that these surfaces are matted, especially in black. Various optical elements can also be used as the transmitted light shielding section 176. For example, it is possible to use filters, mirrors, half mirrors, beam splitters, optical path changing optical elements, and the like. More specifically, various filters such as metals, metal films, and dielectric multilayer films, mirrors and half mirrors, polarizing elements such as polarizing plates and wavelength plates, absorption type ND filters containing light-absorbing substances, metal films, etc. Reflective ND filters, optical path changing optical elements such as various prisms and lenses, etc. may also be used.

The shape and size of the transmitted light from the sample 154 are determined by the shape and incident angle range of the irradiated light 148 that enters the sample cell 152 of the sample section 150 or the sample 154, the structure of the sample cell 152, and the irradiated light 148 in the sample 154. It depends on the spot size, the shape of the condensing lens 142, the focal length f, etc. Therefore, the shape and size of the member used in the parallel scattered light converter 170 are selected depending on the shape and size of the transmitted light 164 from the sample 154. For example, in the case of circular transmitted light, a member of a larger size, such as a disk, is used. When the transmitted light shielding section 176 is arranged between the lens 172 and the scattered light detection section 200, the parallel transmitted light 166 is collimated or diffused by the lens 172, so the arrangement of the transmitted light shielding section 176 Use a member that corresponds to the shape and size of the transmitted light at the position.

The transmitted light blocking section 176 blocks or attenuates the parallel transmitted light 166 from the sample 154. However, in addition to this, scattered light from the sample 154 is also blocked or attenuated. That is, the transmitted light shielding section 176 becomes a shadow object in the measurement of scattered light by the photodetector 202 of the scattered light detection section 200, and the scattering angle corresponding to the shadow portion appearing on the light receiving surface of the scattered light detection section 200 is At this range, measurements of scattered light are difficult. Therefore, it is desirable to accurately measure the parallel small-angle scattered light 168 from the sample 154 to the lower angle side, and for this purpose, the size of the member used for the transmitted light shielding part 176 is made as small as possible. It is preferable that For this reason, it is preferable to use a member for the transmitted light shielding section 176 that is slightly larger than the size of the transmitted light beam to be blocked.

For example, for a sample 154 of latex particles with a diameter of 0.1 μm dispersed in pure water, a 20x infinity-corrected objective lens (f = 10 mm, NA 0.4) is used as the condenser lens 142 of the light irradiation unit 140. As shown in FIG. 2, the sample 154 is placed at a distance LA from the condenser lens 142 to the sample 154 of 10 mm. The irradiation light 148 is made to enter the sample cell 152 at a maximum incident angle θi of 4.3 degrees as shown in FIG. It is placed at a position where the distance LB is 68 mm. Here, the sample cell 152 that stores the sample 154 is constructed by sandwiching a 0.5 mm thick silicone rubber sheet with a 7 mm diameter hole between two 0.15 mm thick microscope cover glasses. are doing. In the above case, the transmitted light 164 from the sample 154 is collimated by 172, and the diameter D1 of the circular parallel transmitted light beam is 10 mm. Therefore, in this case, it is preferable to use, for example, a matte black metal disk or rod with a diameter of 12 mm as the transmitted light shielding part 176.

Further, the effect can be obtained by disposing the transmitted light shielding section 176 either between the parallel scattered light converting section 170 and the scattered light detecting section 200 or between the sample 154 and the parallel scattered light converting section 170. . When the transmitted light shielding section 176 is installed between the sample 154 and the parallel scattered light conversion section 170, the transmitted light from the sample cell 152 of the sample section 150 or the sample 154 is a minute irradiation light beam spot of the sample 154. Since the transmitted light is from the minimum beam spot 157, the size of the member of the transmitted light shielding section 176 can be made smaller than when shielding the parallel transmitted light 166 that has passed through the lens 172. The light shielding section 176 can be made more compact. By making the transmitted light shielding part 176 smaller in size, it becomes possible to measure scattered light at a smaller scattering angle.

Further, the transmitted light shielding section 176 is arranged between the sample section 150 and the scattered light detection section 200 on the optical axis of the transmitted light so as to prevent the transmitted light from entering the photodetector 202 of the scattered light detection section 200. Deploy. Further, the transmitted light shielding section 176 may be placed on the optical axis (traveling direction) of the transmitted light, and there is no restriction on its position (distance) between the sample 154 and the scattered light detection section 200. Note that the transmitted light shielding section 176 does not need to be provided in a configuration that measures rear small-angle scattered light.

(9) Description of the configuration and effects of the scattered light detection section 200 The scattered light detection section 200 includes a photodetector 202 having a light receiving surface 203. Parallel small angle scattered light 168 is detected. The light receiving surface 203 of the photodetector 202 is composed of a large number of photodetecting elements. Each photodetection element 11 outputs an electrical signal corresponding to the light intensity of the incident parallel small-angle scattered light 168, and the electrical signal is amplified by the amplification/cooling section 212 and sent to the analysis section 250.

It is important that the parallel small-angle scattered light 168 is perpendicularly incident on the light receiving surface 203 of the scattered light detection section 200. For this reason, the scattered light detection section 200 has a method for adjusting the light receiving surface 203 of the photodetector 202 to be perpendicular to the optical axis 146, that is, the parallel small angle scattered light 168. The position adjustment mechanism 220 is provided with an angle adjustment mechanism for adjusting the angle of the surface 203 with respect to the optical axis 146. The scattered light detection unit 200 includes an amplification/cooling unit 212 that amplifies the signals from each photodetection element 11 of the photodetector 202 and cools the scattered light detector 202. It includes a control interface (not shown) for electrically transmitting and receiving output signals and control signals to the photodetector 202 and the amplification/cooling unit 212, a driving power source (not shown) for the photodetector 202, etc. are doing.

An example in which the photodetector 202 is used to detect an image of a forward small-angle scattered light image regarding the small-angle scattered light 160 by the scattered light detection unit 200 will be described below. However, the forward small-angle scattered light image based on the small-angle scattered light 160 can be detected in the same way using the light detection camera 204, and the following explanation can be applied as is. That is, the image of the forward small-angle scattered light image based on the small-angle scattered light 160 can be similarly detected using the light detection camera 204 equipped with the camera lens 206 shown in FIGS. 13 and 14. The embodiments shown in FIGS. 13 and 14 specifically describe images of forward small-angle scattered light images created by the action of the ring-shaped aperture 182 of the scattering angle filter 180. However, as described above, it can be similarly used in place of the scattered light detection section 200 in the detection of forward small-angle scattered light images explained using FIGS. 1 to 9, and the same effects can be achieved.

Photodetector 202 is a photodetector suitable for detecting scattered light of sample 154. Specifically, the photodetector 202 is a multi-channel photodetector (consisting of a multi-pixel sensor) including a multi-channel photodiode array, a large number of photosensor element arrays such as a linear image sensor, a CCD image sensor, a CMOS image sensor, etc. equipped with a photodetector). In this embodiment, it is preferable to use a multichannel photodetector as the photodetector 202, which is a large-area photodetector in which a large number of minute photodetection elements are arranged or integrated at high density. In a configuration using a multi-channel photodetector, the intensity distribution of the parallel small-angle scattered light 168 can be measured in a spatial pattern, for example, all at once in a short time while the photodetector 202 is fixed and stationary. In other words, in this case, a scattered light image drawn by scattered light with a multi-channel photodetector is measured as a still image. Further, by continuously photographing the parallel small-angle scattered light 168 using a multi-channel photodetector, in other words, by photographing a moving image, it is possible to measure the temporal change in the spatial pattern described by the intensity distribution, that is, the temporal change.

In this case, the parallel small-angle scattered light 168 is continuously photographed as an image by a multi-channel photodetector, and the obtained moving image of the scattered light images includes the SLS and DLS regarding the sample 154. That is, the positional dependence of the scattered light intensity photographed as an image, that is, the intensity distribution of the scattered light, corresponds to SLS. On the other hand, focusing on a particular point or position in a photographed scattered light image, the time change in the scattered light intensity at that point corresponds to DLS. By continuously photographing scattered light images at certain predetermined time intervals, it is possible to obtain temporal changes in the scattered light intensity at a point of interest on the scattered light image. Based on this acquired data, the analysis unit 250 analyzes the frequency power spectrum or changes in the frequency power spectrum with respect to time changes in the scattered light intensity at each point on the scattered light image, in other words at multiple parts, as necessary. It can be output from.

It is preferable that the photodetector 202 measures the scattered light at once, for example, within a predetermined scattering angle range, for example, in an instant of 100 ms or less, more preferably 50 ms or less. It is preferable that the photodetector 202 can continuously photograph the parallel small-angle scattered light 168 from the parallel scattered light converter 170 as a scattered light image, that is, can take a moving image. As the photodetector 202, for example, a multi-channel photodetector (hereinafter simply referred to as "CMOS camera") including a CMOS image sensor is particularly suitable. With a CMOS camera, for example, continuous shooting is possible at an exposure time of 10 ms, a frame rate of 100 f/s, or 50 f/s, and a continuous shooting length (number of images taken) of 150 to 300 images.

It is preferable that the photodetector 202 of the scattered light detection section 200 has high light reception sensitivity near the light wavelength of the light source 110 used for scattered light measurement. Scattered light from molecules that strongly interact with solvent molecules and their aggregates, such as small to medium molecules such as peptides and nucleic acids, and biopolymers such as proteins, in a solution is dispersed in the liquid and is relatively isolated from the solvent molecules. In general, the interaction is weak compared to nano to submicron particles such as metals, plastics, and inorganic substances and their aggregates. Even the forward small-angle scattered light or the backward small-angle scattered light, which has a relatively large scattering intensity, has an extremely low scattered light intensity compared to the scattered light of particles or aggregates dispersed in the solvent, so the scattered light scattered light It is preferable to use a photodetector with high detection sensitivity as the photodetector 202 of the detection unit 200.

It is necessary that the quantum efficiency of the photodetector 202 itself is high near the light wavelength of the light source 110, and furthermore, the photodetector 202 is used in a highly sensitive state in order to measure weak scattered light. To this end, the light receiving section of the photodetector 202 is cooled in order to suppress dark current noise of the photodetector 202 as much as possible. The light-receiving part of a photodetector can be cooled by air blowing, water circulation, electronic elements, etc. Especially when using an electronic element such as a Peltier element, since the element is small, it is possible to cool the light-receiving part of the photodetector. A device can be built-in. For example, in a photodetector having a quantum efficiency of 50% or more, it is preferable to use a Peltier element to cool the light receiving part of the photodetector to −10° C. or lower to reduce dark current noise.

As the photodetector 202 suitable for the scattered light detection unit 200, a CCD camera having a CCD image sensor or a CMOS camera having a CMOS image sensor, which has a particularly large light receiving surface, is suitable. A CCD camera or CMOS camera for weak light measurement is particularly suitable, with a high quantum efficiency of 50% to 70% or more, very low dark current, large saturation charge, and a linear image sensor with a wide dynamic range. A camera with a side view is preferred. In order to suppress dark current, these cameras may be configured to have a built-in electronic cooler such as a Peltier element in the amplification/cooling section 61 or the like to cool the light-receiving surface to, for example, -10 degrees Celsius or lower. .

As the photodetector 202 of the scattered light detection unit 200, for example, the size of the light receiving unit is 14.0 mm in width x 16.6 mm in height, and the effective pixels are 2160 x 2560 with a minimum pixel size of 6.5 μm (quantum efficiency of 50% or more). A CMOS camera with a 5.5 megapixel CMOS area image sensor in channels (pixels) can be used. Built-in Peltier element allows CMOS sensor to be cooled down to -10℃, and dark current noise is extremely low at 0.02e-/(pixel/second). Also, the dynamic range is wide at 3300:1.

With this CMOS camera, in continuous shooting (video shooting) of scattered light images, etc., for example, if the effective light-receiving surface for shooting is 2160 x 2560 pixels in the entire area, the shooting frame rate is 49 f/s with the global shutter mechanism, and the rolling shutter The mechanism can continuously measure images at 100 f/s. If you want to shoot a video at an even higher speed, for example, set the effective light-receiving surface area to 1080 x 1920 pixels, and shoot the scattered light image etc. at 97 f/s with the global shutter and 200 f/s with the rolling shutter. becomes possible. However, the camera suitable as the photodetector 202 of the scattered light detection section 200 is not limited to the specifications of the CMOS camera described above.

(10) Description of the configuration and effects of the analysis section 250 When the operator inputs measurement conditions for the scattered light detection section 200 from the input/output device 266, the scattered light detection section 200 is controlled based on the input measurement conditions. , the position and angle of the photodetector 202 are set and fixed based on input conditions. After the photodetector 202 is fixed, the intensity distribution of the parallel small-angle scattered light 168 is measured based on the input conditions. If the input condition is to measure a still image, the still image is measured. For example, if the instruction is to shoot a still image multiple times at specified intervals, the shooting will be performed according to the instruction. Moreover, if the input condition is the measurement of a moving image, the intensity distribution of the parallel small-angle scattered light 168 is continuously photographed as a moving image.

The shooting timing and shooting time of still image shooting, moving image shooting, etc. are often controlled according to the passage of time. For this reason, among the control conditions input from the input/output device 266, conditions related to the passage of time are once converted into conditions based on absolute time by the time management device 280 and stored, and the conditions related to the stored time are converted into conditions based on absolute time and stored. Based on the correspondence with the passage of absolute time created by the time management device 280, related configurations of the analysis device 100 are controlled. Control to each component is performed by a control signal 264, which is output from the controller 260 to the related components based on the operation of the CPU 276. Further, measurement results from each configuration and signals representing the state of each configuration are taken into the analysis unit 250 via the controller 260. The same applies to the data measured by the scattered light detection section 200, and the data from the scattered light detection section 200 is taken in from the controller 260 and stored in the storage device 290. The specific analysis operation etc. by the analysis unit 250 will be explained again below.

1.4 Measurement of dynamic light scattering (DLS) and static light scattering (SLS) (1) Overview
As mentioned above, one of the light scattering measurement methods is dynamic light scattering (DLS), which detects light scattering from the sample 154 at a predetermined scattering angle and measures temporal fluctuations in light scattering, in other words, fluctuations. :Dynamic Light Scattering) measurement method. With this DLS measurement method, for example, information regarding the movement of scatterers in the sample 154 to be analyzed can be obtained.

At a predetermined scattering angle θ0, that is, at a certain scattering vector q ₀ , from the time-varying light scattering intensity I(q ₀ ,t), the frequency power spectrum I(ω) and the correlation can be calculated by Fourier analysis and correlation function analysis with respect to time. A function G(τ) and a scattering factor S(q ₀ , ω) are obtained. Here, ω is the angular frequency and τ is the relaxation time.

Furthermore, in the analysis of the DLS measurement method, the diffusion coefficient D of the scatterer is often calculated from the correlation function G(τ) to evaluate the size of the scatterer, that is, the value of the hydrodynamic particle size Rh. Here, the particle size Rh of the scatterer evaluated from the diffusion coefficient D is valid only when the scatterer model such as spherical is applicable to the analysis target such as fine particles or aggregates. It is necessary to keep this in mind. In addition, to calculate Rh, it is necessary to use the measured value of the viscosity coefficient (viscosity) η and temperature T of the sample solution containing fine particles and aggregates, and in particular, the value of the viscosity coefficient η apart from the DLS measurement. will be measured.

Another light scattering measurement method is a method of measuring the dependence of the scattering angle on the average intensity of light scattering from an analysis target, and is a static light scattering (SLS) measurement method. The SLS measurement method mainly provides information regarding the structure of the scatterer of the sample 154 to be analyzed.

The average light scattering intensity I(q,ω ₀ ) averaged over a certain time t ₀ is measured at various scattering angles θ (scattering vector q), and the structure factor S(q,ω ₀ ) inside the scatterer is obtained. It will be done. Here, ω ₀ is the angular frequency with respect to t ₀ . In the SLS measurement method, the intensity of scattered light in samples of different concentrations is often measured in dilute solutions such as polymers or colloids, where the concentration of scatterers is sufficiently low that interference effects between scatterers can be ignored. In the SLS analysis, molar mass (molecular weight), radius of gyration, and second virial coefficient are analyzed and evaluated from, for example, Zim plot, Berry plot, etc.

In addition, in the SLS measurement method, the scattering angle dependence of the average light scattering intensity is measured, particularly at a low angle forward or backward, and the fractal dimension, which is an index of the structural parameter (structural factor) of the scatterer, is analyzed and evaluated. That _is , the _structure factor S(q , ω ₀ ) is the spatial Fourier transform of the average local density function, so that factors directly related to the fine structure of the scatterer can be evaluated.

That is, the average intensity I(q,ω ₀ ) of forward scattered light or backward scattered light measured for particles or aggregates exhibits a power function type. That is, it is expressed as I(q,ω ₀ )=I ₀ q ^−α . Here, _I0 is a constant, α is a power, and a measurement object in a three-dimensional space has a value of 0<α<3. Then, from curve fitting using a power function of the measured average intensity I(q,ω ₀ ), the power number α is evaluated, and the fractal dimension Df of the aggregate or agglomerate of the measured particles is evaluated.

Conventionally, in the analysis of particles by light scattering measurement, the DLS measurement method has been used to analyze and evaluate particle size and its distribution, while the SLS measurement method has been used to analyze and evaluate molecular weight and the like. That is, depending on the purpose of analysis and evaluation, analyzers employing one of the light scattering measurement methods have been used separately. There has not been an analyzer that can measure both the DLS measurement method and the SLS measurement method and perform both types of analysis using the same analyzer.

Furthermore, the DLS measurement method and the SLS measurement method are not only very different in the method of measuring light scattering as described above, but also in the method of analyzing the measurement data. If two characteristic aspects of light scattering, i.e., temporal measurement (DLS measurement) and spatial measurement (SLS measurement) of light scattering can be simultaneously measured for the same analysis target, it is possible to analyze targets such as particles and aggregates. It is possible to simultaneously analyze and evaluate the knowledge regarding the motion and structure of the object using a single light scattering measurement device, which has a very large effect.

Furthermore, micron-order droplet formation due to liquid-liquid phase separation, which has recently attracted particular attention in polymer gelation, crystallization of proteins, peptides, etc., and intracellular metabolic reactions, and non-uniformity in solutions. There is a great need for microstructural analysis and evaluation of solutions on a submicron microscopic scale to deal with the aggregation of proteins, peptides, etc. that occurs in microscopic systems. The analyzer 100 of the embodiment can fully meet these needs.

(2) Description of scattered light detection in the analyzer 100 of the embodiment As described above, in the analyzer 100, the DLS measurement method and the SLS measurement method are performed using the scattered light detection unit 200 on the sample 154 that is the target of analysis. It is possible to obtain measurement data that can be used for both. Before explaining the configuration for measurement, the results measured by the configuration of the analyzer 100 will be explained using FIGS. 4 and 5.

FIG. 4 is a captured image 230 obtained by measuring and analyzing parallel small-angle scattered light 168 based on small-angle scattered light 160 generated from the minimum beam spot 157 by the photodetector 202 of the scattered light detection unit 200. A small-angle scattered light image by small-angle scattered light 160 from the minimum beam spot 157 is magnified on a microscopic scale and displayed as a microscopic image on the photodetector 202 of the scattered light detection unit 200, and this microscopic image is stored in the storage device 290 as a measurement result. This is a photographed image created based on the data of the stored microscopic image that is the measurement result. On the side closer to the optical axis 146 is a dark area 232 that is generated because the small-angle scattered light from the sample 154 is blocked by the transmitted light blocking section 176. An image portion 234 based on the measurement of the parallel small-angle scattered light 168 extends on the outer circumferential side of the dark portion 232 .

(3) Explanation of the analysis of the DLS analysis region R1 in the image section 234 In FIG. 4, the measurement data of the small-angle scattered light 160 in the DLS analysis region R1 is calculated over a predetermined period of time when analysis based on the DLS measurement method is performed. Measurements are repeated each time. In this case, in this embodiment, measurement data related to the DLS analysis region R1 obtained through repeated measurements can be specified, and the DLS measurement method can be implemented.

For example, when the operator specifies the DLS analysis region R1 as an analysis target by the DLS measurement method, the analysis unit 250 determines that the coordinates of the DLS analysis region R1 are the coordinates (xp, yp), as shown in FIG. It is obtained from the operation in . The analysis unit 250 can specify data regarding the DLS analysis region R1 at each measurement time of the X coordinate value xp and the Y coordinate value yp in the determined coordinates (xp, yp). Therefore, by reading the data at each measurement time of the DLS analysis region R1 from the measurement data obtained by continuously repeating measurements at predetermined time intervals, and displaying the changes as a graph with time as a parameter, the DLS Analysis results based on the measurement method can be output from the input/output device 266.

(4) Description of method for specifying measurement data regarding DLS analysis region R1 FIG. 6 is a partially enlarged view of the image detection surface of an example of the photodetector 202 in the scattered light detection section 200. Each photodetection element is arranged to form a two-dimensional plane, for example, photodetection element 11 to photodetection element 12, photodetection element 13, etc. are arranged in the X-axis direction, and in the Y-axis direction. A photodetection element 11, a photodetection element 21, a photodetection element 31, and so on are arranged. According to the X-axis direction and the Y-axis direction, the detection results of each photodetection element are transmitted from the scattered light detection section 200 to the analysis section 250 in a predetermined order, and are stored in the storage device 290 of the analysis section 250, for example, in the order of addresses. is retained. The order of addresses held in the storage device 290 has a predetermined correspondence with each photodetector element. Therefore, the measurement data of the photodetector element to be used can be read by specifying the address of the storage device 290. Therefore, the image based on the small-angle scattered light 160 detected by the photodetector 202 is divided finely in the two dimensions of the X axis and the Y axis, and the detection data for each divided area is stored in the storage device 290 for each of the divided areas. The area and the storage position in the storage device 290 are stored while maintaining a correlation with each other.

Furthermore, under the conditions set by the operator, the photodetector 202 maintains the image detection operation based on the small-angle scattered light 160, and detects the image for each area, in other words, from the photodetector elements 11 to 34. The detection results for each area are repeatedly stored in the storage device 290 under the above conditions. That is, temporal changes in the image based on the small-angle scattered light 160 are stored in the storage device 290. Based on the detection results stored in the storage device 290, it is possible to analyze changes in measurement results over time for the set area, and the analysis unit 250 can perform DLS analysis.

In addition, the forward small-angle scattered light image detected by the photodetector 202 includes an area corresponding to the optical axis 146 and a plane perpendicular to the optical axis 146, as shown in FIGS. 4 and 5. The area related to the radial image in the area is included. The forward small-angle scattered light image of a region extending in the radial direction from the optical axis 146 including the region designated by the operator is divided into fine sections, measured for each of the finely divided measurement unit areas, and the measurement results are divided into the finely divided areas. The location of each measurement unit area is stored in the storage device 290 in a state where the location can be specified. Therefore, the detection results necessary for SLS analysis in the specified area can be read out and used, thereby enabling SLS analysis related to the specified area.

When the positional relationship between the optical axis 146 and the photodetector 202 of the scattered light detection unit 200 is specified, the coordinates (x0, y0) of the optical axis 146 and the DLS analysis region R1 coordinates (xp, yp) shown in FIG. ) and the photodetecting element shown in FIG. 6 are in a state of being associated with each other. Therefore, the address of the storage device 290 where the detected data is held is determined from the associated photodetection element, and the measurement results of the coordinates (x0, y0) and DLS analysis area R1 coordinates (xp, yp) to be used are transferred to the wiring. 205. Here, each of the photodetecting elements corresponds to an area of each measurement unit described above.

When performing analysis based on the DLS measurement method, it is necessary to store data measured at predetermined time intervals in the DLS analysis region R1 each time, and to read out data at different measurement points stored when performing analysis. Become. FIG. 7 shows a state in which data measured at predetermined time intervals is stored. When measuring N times at measurement time points T1, T2...TN, for example, in order to store the measurement results for each measurement time point T1, T2...TN, blocks B1, Secure locks B2, . . ., TN. The detection results of each photodetecting element of the photodetector 202 at the measurement time T1 are stored in the address associated with each photodetecting element in the area of block B1 of the storage device 290. Similarly, the detection results of each photodetecting element of the photodetector 202 at measurement time T2 are stored in the address associated with each photodetecting element in the area of block B2. By repeating this N times, it is possible to store the detection results of each photodetection element of the photodetector 202 by associating the detection results up to the measurement time TN with the addresses in the associated blocks.

When performing analysis based on the DLS measurement method in the DLS analysis region R1, measurement data of the DLS analysis region R1 at different detection times T1...TN can be read from each block B1...BN of the storage device 290. , it becomes possible to perform analysis based on the DLS measurement method from these measurement data.

It is very important to set the DLS analysis region R1 at the desired position for analysis. By creating and displaying the captured image 230 shown in FIG. 4 in the analysis unit 250 and setting the DLS analysis region R1 based on the captured image 230, a very useful DLS analysis result can be obtained. By the method described above, the DLS analysis region R1 is set, for example, on the screen of the photographed image 230. By the calculation of the analysis unit 250, the photodetecting element in the photodetector 202 corresponding to the DLS analysis region R1 is specified, and data corresponding to the DLS analysis region R1 is read out for each block in FIG. DLS analysis becomes possible for the area to be analyzed.

(5) Description of method for specifying measurement data regarding SLS analysis region R2 Next, the SLS measurement method in the analyzer 100 will be described using FIGS. 8 and 9. The photographed image shown in FIG. 8 is the photographed image previously explained using FIG. This is a captured image obtained by measuring and analyzing parallel small-angle scattered light 168 based on . The SLS measurement method is a measurement method that measures the scattering angle dependence of the light scattering intensity at a low angle, and will now be explained by taking as an example the measurement of the SLS analysis region R2 shown in FIG. 8. The parallel small-angle scattered light 168 is measured by the photodetector 202 shown in FIG. In FIG. 8, a dark portion 232 of the small-angle scattered light 160 near the optical axis 146 is a portion where the parallel small-angle scattered light 168 and the parallel transmitted light 166 are blocked by the transmitted light shielding portion 176. The image portion 234 represents the intensity of the small angle scattered light depending on the scattering angle with the optical axis 146 of the parallel small angle scattered light 168 as a reference.

When the SLS analysis region R2 in the image section 234 is specified, the photodetecting element of the photodetector 202 corresponding to the SLS analysis region R2 is specified, as described above. The photodetecting elements of the photodetector 202 corresponding to the X-axis direction of the SLS analysis region R2 are, for example, the photodetecting elements 21 to 2N shown in FIG. Further, the photodetecting elements corresponding to the width of the SLS analysis region R2 in the Y-axis direction are the photodetecting element 11, the photodetecting element 21, and the detecting element 31. In reality, the width of the SLS analysis region R2 in the Y-axis direction is wider than the photodetection element 21 and the detection element 2N, and there is a possibility that a plurality or many photodetection elements are included in the Y-axis direction. In that case, the average value of the measurement results of the photodetecting elements included in the width in the Y-axis direction may be used.

In the above description, the SLS analysis region R2 was set in the X-axis direction for the sake of simplicity. However, when determining the scattering angle, an angle other than the X-axis direction can be freely determined. For example, the image section 234 shown in FIGS. 4 and 8 is displayed by the analysis section 250, and the operator can set the SLS analysis region R2 based on the image section 234 shown in FIGS. 4 and 8. When the SLS analysis region R2 is set, the photodetection element in the photodetector 202 corresponding to the set SLS analysis region R2 is determined, and the measured values of the corresponding photodetection element of the photodetector 202 are shown in FIG. It becomes possible to read from the address of the storage device 290 described. By analyzing the measurement data read from the address of the storage device 290 according to the distance from the photodetection element corresponding to the optical axis 146, the analysis result of the SLS analysis region R2 can be obtained.

2. Description of Other Embodiments (1) Outline of Other Embodiments As other embodiments, the analyzer 102 in which functions are added or modified with respect to the embodiments shown in FIGS. 1 to 9 is shown in FIGS. 14. In the configuration of the analyzer 102 shown in FIG. 10, a light source 110, an incident light adjustment section 120, an optical path changing section 130, an optical aperture 138, a light irradiation section 140, a sample section 150, a parallel scattered light conversion section 170, a scattered light detection section 200, the analysis section 250, and the like have already been described as the configuration of the analysis device 100. Structures with the same reference numerals as those shown in FIGS. 1 to 9 are the same structures, and have the same effects, unless a particular explanation is added. The major difference between the analyzer 102 and the analyzer 100 is that a scattering angle filter 180 and a scattered light condenser 190 are provided between the parallel scattered light converter 170 and the scattered light detector 200. Differences in analysis content and points for improvement due to this will be explained below.

(2) Description of the scattering angle filter 180 and the scattered light condenser 190 The scattering angle filter 180 and the scattered light condenser 190 will be explained using FIGS. 11 and 12. A scattering angle filter 180 having an aperture 182 is arranged between the parallel scattered light converter 170 and the scattered light condenser 190. The specific angle parallel small angle scattered light at the selected angle is selected, and the specific angle parallel small angle scattered light at the selected angle enters the scattered light condensing unit 190 as arc-shaped specific angle scattered light 186 described below. The incident arc-shaped specific angle scattered light 186 is converted into small-angle scattered convergent light 196 by the scattered light condenser 190, guided to the photodetector 202 of the scattered light detector 200, and its intensity etc. are measured.

The scattered light condensing unit 190 includes a lens 192, a support holder 194, and the like. The support holder 194 holds the lens 192 and includes a mechanism for adjusting the position and tilt angle of the lens 192. Further, the scattering angle filter 180 transmits light forming an arc shape centered on the optical axis 146 whose radius is the length corresponding to the set angle, out of the parallel small angle scattered light 168. This arc-shaped light selected by the scattering angle filter 180 is shown as arc-shaped specific angle scattered light 186 in FIGS. 10 and 11, and FIGS. 13 and 14. The scattering angle filter 180 has a function of selecting arcuate specific angle scattered light 186 from the parallel small angle scattered light 168 and transmitting it. In this embodiment, the scattering angle filter 180 also has the function of the transmitted light blocking section 176 in the previous embodiment, which prevents the parallel transmitted light 166 from the sample 154 from entering the scattered light detection section 200. .

As shown in FIG. 12, the scattering angle filter 180 includes a light shielding plate 181 in which a ring-shaped opening 182 is formed. The parallel transmitted light 166 based on the transmitted light 164 from the sample 154 is blocked by the light shielding plate 181 and can be prevented from entering the photodetector 200. The light shielding plate 181 is arranged perpendicularly to the optical axis 146, and the light shielding plate 181 transmits only light having a specific radius in a plane perpendicular to the optical axis 146. Since the specific radius corresponds to the specific scattering angle of the scattered light, it is possible to selectively transmit only the specific-angle parallel small-angle scattered light of the set specific scattering angle. As will be explained below, the aperture 182 formed in the scattering angle filter 180 has an arcuate shape that forms at least a part of the circumference with a radius determined by the specific angle with respect to the optical axis 146. . Therefore, the light transmitted through the aperture 182 of the scattering angle filter 180 has an arc shape, and the arc-shaped specific angle scattered light 186 is selected by the scattering angle filter 180 out of the parallel small angle scattered light 168. The scattered light passes through 180 and enters the scattered light condenser 190 .

In FIG. 12, the light shielding plate 181 includes a circular inner light shielding part 183 centered on the optical axis 146, an opening 182, and a peripheral light shielding part 185. The arcuate specific angle scattered light 186 having the set scattering angle is selectively passed through the aperture 182. Therefore, the inner light shielding part 183 provided on the optical axis 146 side of the ring-shaped opening 182 blocks the parallel transmitted light 166 inside a specific scattering angle, and the peripheral light shielding part 185 blocks the parallel small-angle scattered light 168. The parallel small-angle scattered light 168 on the outer peripheral side of the opening 182 is blocked. As shown in FIG. 12, the scattering angle filter 180 forms a ring-shaped opening 182 and has a plurality of bridges 187 connecting the peripheral light shielding part 185 and the inner light shielding part 183 in order to support the inner light shielding part 183. ing.

As described above, the aperture 182, the inner light shielding part 183, and the surrounding light shielding part 185 of the scattering angle filter 180 are configured according to the shape of the parallel small angle scattered light 168, the shape of the arcuate specific angle scattered light 186 at the selected scattering angle, and its selection. It is determined depending on the range of the scattering angle, the shape of the parallel transmitted light 166 to be blocked, the shape of the photodetector 202 of the scattered light detection section 200, etc. When the parallel small-angle scattered light 168 is a circular image, the shape of the aperture 182 of the scattering angle filter 180 is preferably ring-shaped. For example, when the parallel small-angle scattered light 168 is a rectangular image, the shape of the aperture 182 of the scattering angle filter 180 is preferably a linear slit.

The material of the scattering angle filter 180 is not particularly limited as long as it does not allow the transmitted light 164 from the sample 154 of the sample section 150 to pass through. For example, various metals, plastics, wood, etc. can be used. Furthermore, it is desirable that the transmitted light 164 from the sample 154 be irradiated onto the scattering angle filter 180 to prevent stray light from occurring. For example, it is preferable to apply a matte black coating to the surface of the scattering angle filter 180, particularly the side irradiated with the transmitted light 164.

Specifically, for the scattering angle filter 180, as shown in FIG. A ring-shaped opening 182 with a gap of 1.0 mm is formed between the inner light shielding part 183 and the circular peripheral light shielding part 185. This scattering angle filter 180 having a ring-shaped aperture 182 of 1.0 mm is used as the lens 172 of the parallel scattered light converter 170, for example, as a 20x infinity-corrected objective lens (focal length f is 10 mm, numerical aperture NA is 0.4) and the distance LB from the sample 154 to the objective lens 172 of the parallel scattered light converter 170 is set to 15 mm, for example, the parallel small angle scattered light 168 is calculated as the radiation from the sample 154. The angle θ0S (see FIG. 3) is selected in the scattering angle range of 16.7 degrees to 20.2 degrees so that the light can pass through the ring-shaped aperture 182 of this scattering angle filter 180. In addition, when the beam diameter of the transmitted light 164 from the sample 154 by the objective lens 172 of the parallel scattered light conversion unit 170 is, for example, a radius of 1.5 mm, the maximum angle θ0S (see FIG. 3) is calculated as the maximum angle Since θ0S is 5.7 degrees, the parallel small-angle scattered light whose measured scattering angle θS is in the range of 11.0 degrees to 14.5 degrees is selected by the scattering angle filter 180 and scattered. The light is introduced into the lens 192 of the light condensing section 190.

The parallel small-angle scattered light in a predetermined narrow scattering angle range selected by the scattering angle filter 180 is focused by the lens 192 of the scattered light condensing section 190 and is collected as a scattered light image by the photodetector 202 of the scattered light detection section 200. Detected. Therefore, since the focused scattered light image detected by the scattered light detection unit 200 is the arc-shaped specific angle scattered light 186 that has passed through the aperture 182 of the scattering angle filter 180, the shape of the aperture 182 of the scattering angle filter 180 is The image will be reduced or enlarged. Note that in this embodiment, the aperture 182 has a nearly circular shape in order to produce an ideal image. However, if the aperture 182 is a circular arc, an image corresponding to the circular arc can be created. For example, even if the arc is about one-sixth of the circumference, an image corresponding to the arc is generated. However, if the aperture is a point, the amount of light passing through it will be so insufficient that virtually no image will result.

The arc-shaped specific angle scattered light 186 that passes through the scattering angle filter 180 and enters the lens 192 of the scattered light concentrator 190 is collimated by the lens 172 of the parallel scattered light converter 170. In particular, for example, when the focal position of the lens 192 of the scattered light condensing section 190, that is, the distance L1 between the lens 192 and the light receiving surface of the scattered light detection section 200, is arranged equal to the focal length of the lens 192, The focused scattered light image of angularly scattered light 186 has a minimum size. In particular, if it is assumed that the scattered light is emitted from an infinitesimal point on the sample 154, the scattered light image will theoretically also be an infinitesimal point.

However, in reality, as described above, the irradiation light 148 that enters the sample 154 has a finite and minute diameter, for example, a circular incident beam of several μm to several tens of μm. Even if the distance from the light-receiving surface of the lens 168 is set equal to the focal length of the lens 192, the focused scattered light image of the parallel small-angle scattered light 168 is not a dot-like image but a finite-sized circular image. It is detected by the detection unit 200.

Furthermore, when the light receiving surface of the scattered light detection section 200 is arranged before and after the focal point of the lens 192 of the scattered light condensing section 190, the parallel small angle scattered light 168 that passes through the scattering angle filter 180 and is focused by the lens 192. The scattered light image is an image in which the shape of the aperture 182 of the scattering angle filter 180 is projected onto the light receiving surface of the scattered light detection unit 200, and the light image that has passed through the aperture 182 is reduced or expanded.

Irradiation light 148 formed by the condensing lens 142 of the light irradiation unit 140 enters the sample 154, and scattered light is emitted from the minimum beam spot 157 (see FIG. 3), which is a minute irradiation light area. Scattered light emitted from two scatterers located at slightly different positions within the irradiated light area of the sample 154, for example, two scatterers separated by a few μm or less in the sample 154, is emitted from slightly different positions, so that the scattered light is slightly different. Follow different optical paths. Therefore, the optical path of those scattered lights is changed by the lens 172 of the parallel scattered light converter 170, a part of it is selected by the scattering angle filter 180, and the optical path is changed by the lens 192 of the scattered light condenser 190. A scattered light image is formed.

The scattered light image detected by the scattered light detection unit 200 is formed including the following two types of scattered light. That is, one is that scattered light is emitted by a scatterer from an irradiated light area that can be considered as a minute point on the sample 154, and the scattered light is parallelized (collimated) by the lens 172 of the parallel scattered light conversion unit 170 and converted into parallel light. This becomes small-angle scattered light 168. A part of the scattered light within a certain predetermined scattering angle range is selected from the parallel small-angle scattered light 168 by the scattering angle filter 180, and is a scattered light image formed by the small-angle scattered convergent light 196 that is focused by the lens 192.

The other is that scattered light is emitted from a group of scatterers at different positions within the irradiation light region that can be considered to be of finite size in the sample 154, and the optical path of the scattered light is changed by the lens 172 of the parallel scattered light converter 170. , these scattered lights are substantially collimated by the lens 172. A scattered light image is formed by selecting some of the scattered light by the scattering angle filter 180 and changing the optical path of the scattered light by the lens 192. Therefore, the scattered light image formed by the latter scattered light reflects the positional distribution of the scatterers within the irradiated light area in the sample 154. That is, the scattered light image formed by the latter scattered light represents a microscopic image of the sample 154 that is a microscopic image of the scattered light.

The scattered light images formed by the above-mentioned two types of forward small-angle scattered light and backward small-angle scattered light on the measurement sample 154 follow different optical paths and are imaged, so that they are detected by the scattered light detection unit 200. The shape of the opening of the scattering angle filter 180 appears at a different position from the projected scattered light image. Specifically, for example, when using the scattering angle filter 180 having a ring-shaped opening as described above, the scattering angle filter is formed by parallel forward small-angle scattered light or parallel backward small-angle scattered light focused by the lens 192. A bright ring-shaped scattered light image onto which the ring-shaped apertures 180 are projected appears near the center. A scattered light image reflecting the position distribution appears. That is, the microscopic image of the scattered light appears in a dark field area different from the bright scattered light image projected by the ring-shaped opening of the scattering angle filter 180. Therefore, for example, with the configuration shown in FIG. 11, simultaneous measurement of dynamic and static forward small-angle scattered light can be performed on the sample 154 on a microscope scale.

Note that in FIG. 11, the scattered light detection section 200 has a photodetector 202 for detecting a microscopic image. In the embodiment shown in FIG. 13, a light detection camera 204 including a translucent screen 208 and a camera lens 206 is used as the scattered light detection section 200. Further, in the embodiment shown in FIG. 14, a light detection camera 204 equipped with a camera lens 206 is used as the scattered light detection unit 200. The detection results of the light detection camera 204 are similar to the operations and effects of the scattered light detection section 200 described above. The measurement results are taken into the analysis unit 250 and stored in the storage device 290, and the stored results are used for DLS analysis and SLS analysis.

In the example of use of the photodetector 202 described above, a microscopic image by the parallel small-angle scattered light 168 is created on the measurement surface of the photodetector 202, and the image is segmented and measured by the photodetector 202. The same applies to the light detection camera 204 shown in FIGS. 13 and 14, in which the microscopic image created by the small-angle scattered convergent light 196 taken in through the camera lens 206 is divided into subdivided areas and sequentially displayed. The data is captured and stored in the storage device 290.

In the embodiment shown in FIG. 13, an image based on small-angle scattered convergent light 196 is displayed on a screen, a microscope image displayed on the screen is photographed by a light detection camera 204 equipped with a camera lens 206, and the photographed microscope image is The image is stored in the storage device 290 of the analysis unit 250. In FIG. 13, a translucent screen 208 is used as the screen. By doing so, the microscope image can be taken from the rear side of the semi-transparent screen 208, which is the back side of the semi-transparent screen 208. In this case, it is easy to arrange the camera lens 206 of the photodetection camera 204 in alignment with the optical axis 146, and there is an effect that the entire device can be made smaller.

Further, the embodiment shown in FIG. 14 is an example in which the camera lens 206 of the light detection camera 204 is arranged behind the focal point position of the small-angle scattered convergent light 196. The size of the microscopic image created by the small-angle scattered convergent light 196 can be changed depending on whether it is placed behind or in front of the focal position of the small-angle scattered convergent light 196.

Note that in FIGS. 13 and 14, the amplification/cooling unit 212 is a cooling device for amplifying the photographed signal and lowering the level of electrical noise generated inside the photodetection camera 204. It functions similarly to the amplification/cooling section 212 shown in FIG. 2 and produces effects.

3 Description of Analysis Unit 250 3.1 Configuration of Analysis Unit 250 The analysis unit 250 includes a controller 260, an input/output device 266, a CPU 276, a storage device 290, and the like. The analysis unit 250 controls the overall operation of the analysis device 100 or 102, and also analyzes the measurement results of the scattered light detection unit 200 equipped with the photodetector 202 or the light detection camera 204 that operates as the scattered light detection unit 200. is taken in, DLS analysis and SLS analysis are performed, and the results are output from the input/output device 266. A controller 260 is provided to control each component of the analyzer 100 and the analyzer 102, and control signals are sent from the controller 260 to each component in response to instructions from the CPU 276. Furthermore, the measurement results from the photodetector 202 of the scattered light detection section 200 and the light detection camera 204 that operates as the scattered light detection section 200 are taken in via the controller 260 and recorded in the storage device 290 as described above. .

When the operator inputs instructions, the instructions are input from the input/output device 266. Further, measurement results and analysis results are output from the input/output device 266. The input/output device 266 includes a keyboard, a display device, a printing device, etc. in order to perform these operations.

3.2 Description of the operation of the analysis unit 250 FIG. 15 is an explanatory diagram illustrating an example of the operation of the analyzer 100 and the analyzer 102. For example, in this embodiment, before entering the measurement analysis mode M2, which imports measurement results for DLS analysis and SLS analysis, and executes DLS analysis and SLS analysis based on the imported measurement results, adjustment mode M1 is activated. An action is taken. Adjustment mode M1 for adjusting the position of sample cell 152 by sample position adjustment mechanism 158 of sample section 150 is started when an instruction to start adjustment mode M1 is input from input/output device 266. In the adjustment mode M1, the position of the sample cell 152 is adjusted by the sample position adjustment mechanism 158 so that the irradiation position of the irradiation light 148 matches the position to be analyzed. In reality, an image of the small-angle scattered light 160 from the current irradiation position of the irradiation light 148 is captured by the analysis unit 250 from the scattered light detection unit 200 or the light detection camera 204 that operates as the scattered light detection unit 200. The above image is displayed on the input/output device 266 as an adjustment image. The operator can adjust the position of the sample cell 152 using the sample position adjustment mechanism 158 while viewing the adjustment image that is displayed on the input/output device 266. When the position to be analyzed is found based on the image displayed on the input/output device 266 and the setting of the position of the sample cell 152 by the sample position adjustment mechanism 158 is completed, the adjustment mode M1 ends. The control and operation of the analysis unit 250 for this termination may be performed by the operator's instruction to terminate, or may be performed by the operator's instruction to start the measurement analysis mode M2.

In the adjustment mode M1, an image based on data taken in from the scattered light detector 200 or the photodetector 202 is displayed on the input/output device 266 as an adjustment image. However, calculations for DLS analysis and SLS analysis based on the detection results of the scattered light detection unit 200 and the light detection camera 204 used as the scattered light detection unit 200 are not executed. Further, the continuous and repeated loading operation of the measurement results of the scattered light detection section 200 or the light detection camera 204 used as the scattered light detection section 200 into the storage device 290 is also stopped.

Note that the light detection camera 204 used as the scattered light detection unit 200 and the scattered light detection unit 200 is experimentally determined based on the small-angle scattered light 160 from the irradiation position of the irradiation light 148 determined by the current setting position of the sample cell 152. The measurement results may be repeatedly taken in on a trial basis, the results may be used to perform a trial DLS analysis or SLS analysis, and the position of the sample cell 152 may be readjusted based on the results. In this embodiment, the above-described experimental DLS analysis and SLS analysis may be performed in the measurement analysis mode M2 described below, and the sample cell 152 may be readjusted by the sample position adjustment mechanism 158 based on the results. It is possible.

After the adjustment of the positioning of the sample cell 152 in the adjustment mode M1 is completed, an operator's instruction to start measurement is input from the input/output device 266, and the measurement analysis mode M2 is started. An example of the measurement analysis mode M2 will be explained with reference to FIG. Based on the initial setting described later, the number of times of photographing is 1 to the number of times of photographing is N, that is, the measurement results from the scattered light detection unit 200 and the photodetector 202 are taken. In order to perform DLS analysis or to shoot a video during the number of shooting times 1, the measurement results of the scattered light detection unit 200 and the light detection camera 204 that operates as the scattered light detection unit 200 are repeatedly and continuously captured and stored. It is held in the device 290. Here, one time of capturing measurement results from the scattered light detection unit 200 or the photodetector 202 is referred to as sampling.

When the imaging time T1 for each of the imaging times 1 to N is set, the above-mentioned steps are repeated at the set sampling frequency as shown in FIG. 15 within the imaging time T1 assigned to each imaging number. The measurement results are continuously loaded into the storage device 290. The sampling execution period is determined by the sampling frequency input in the initial settings. The number of samplings at the imaging time T1, which is the number of times of repeated loading into the storage device 290, may be set by directly inputting the number of samplings, or may be set by inputting the duration of the repetition, which is the imaging time T1. good. The sampling operation is performed for the set N times in the set shooting time T1, or is performed for the set shooting time T1 which is the duration of continuous acquisition of the measurement results, etc. The sampling operation based on this is continuously repeated. When the photographing time T1 of the sampling operation ends, the operation described in FIG. 15 ends, for example, when the number of times of photographing is 1.

The sampling operation is stopped during the set imaging interval T2. The time management device 280 of the analysis unit 250 has a function to manage the start and end timings of control operations, and when the shooting interval T2 ends, the CPU 276 starts shooting based on the report from the time management device 280. Issue an instruction to start the second operation. The operation described above as the operation for the number of times of photographing 1 is performed as the number of times of photographing is 2. Thereafter, the sampling operation is stopped again during the imaging interval T2.

When the above-mentioned operation is repeated and the execution of the number N of images is completed, the measurement analysis mode M2 is ended, and the acquisition of data for DLS analysis and SLS analysis is completed. Based on this data, calculations for DLS analysis and SLS analysis are performed, and the analysis results are output from the input/output device 266. Note that the method of terminating the operation of the measurement analysis mode M2 by repeating the number of photographing operations described above N times is one example. That is, instead of setting the number N of the number of times of photographing, a method may be used in which the total photographing time TA for photographing is set. The conditions for the photographing time T1 and the subsequent photographing interval T2 in which the operation of continuously and repeatedly capturing the measurement results of the scattered light detection unit 200 is continued are the same, but the repetition of the photographing time T1 and the subsequent photographing interval T2 is not limited to the number of repetitions. , the total photographing time TA, and it is also possible to set this total photographing time TA as the initial setting. In this case, after the total imaging time TA has elapsed, the operation of capturing the measurement results of the scattered light detection unit 200 based on the new imaging time T1 is not performed. Thereby, it is possible to control the end of the measurement analysis mode M2.

The operation shown in FIG. 15 describes the acquisition of measurement results from the scattered light detection unit 200 and the light detection camera 204, which is another specific example of the scattered light detection unit 200. The content shown in FIG. 15 is an example, and changes depending on the instruction content of the operator. In addition, it is possible to perform some calculations for DLS analysis and SLS analysis based on the captured data without waiting for the end of the number of shots N. That is, in parallel with the operation of capturing the measurement results of the scattered light detection unit 200 in the measurement analysis mode M2, calculations for the above-mentioned DLS measurement method or SLS measurement method can be performed based on the captured measurement results.

FIG. 16 is a flowchart for the CPU 276 to execute the operation of the measurement analysis mode M2 shown in FIG. FIG. 17 is an explanatory diagram illustrating the storage state of measurement results in the storage device 290 and the storage state of analysis results using the stored data. FIG. 16 shows the operation in the measurement analysis mode M2, which is a state in which the adjustment mode M1 has already ended. When the operation is started in step S300, the above-mentioned initial settings are performed by the operator in step S302. In the initial settings, for example, the imaging time T1 for each imaging number from 1 to N, the sampling frequency executed for each imaging from 1 to N, and the time interval of intermittent imaging. The setting contents shown in FIG. 15, such as T2, the total number of repetitions N of the number of imaging times, the DLS analysis area, the SLS analysis area, and the total imaging time TA, are input from the input/output device 266. Thereafter, the execution of the CPU 276 moves to step S304, and the measurement results from the scattered light detection unit 200 at the number of shots 1 shown in FIG. 15 are taken in, that is, the measurement results from sampling 1 to sampling N shown in FIG. 15 are taken in. is executed and stored in memory block B1 shown in FIG.

When the continuous operation of importing the measurement results for the number of shots 1 is completed, step S310 is executed, and the measurement results corresponding to the DLS analysis area and the SLS analysis area are selectively extracted from the measurement results stored in the memory block B1. The data is stored in addresses secured as DD1 and SD1 of the storage device 290 shown in FIG. Next, in step S312, a calculation for static scattered light analysis is executed based on the data stored at the address SD1, and in step S314, the calculation result is stored in the address ARS1 of the storage device 290.

Further, in step S322, calculations for analyzing dynamic scattered light are executed, and the results are stored in address ARD10 and address ARD11 in step S324. After step S324 is executed, it is determined in step S330 whether the execution is finished, and based on the completion of the number of shots 1, the operation related to the next number of shots 2 is started in step S304. Although the operation in step S304 starts after the photographing interval T2 has elapsed, the flowchart of FIG. 16 omits a description of the passage of the photographing interval T2. Actually, the time management device 280 described above manages time, performs interrupt processing, etc., controls the timing of starting the operation for the number of times of shooting 2, and starts the operation in step S304.

Instead of setting the execution time of the measurement analysis mode M2, that is, the end condition of the measurement analysis mode M2, with the number of shots 1 to the number N of shots, when the end condition of the measurement analysis mode M2 is set with the total shooting time TA, In step S330, it is determined whether the time elapsed in the measurement analysis mode M2 has reached the total imaging time TA based on the time elapsed in the measurement analysis mode M2, and if the time elapsed in the measurement analysis mode M2 has reached the total imaging time TA. In this case, instead of starting execution of step S304, the execution of the CPU 276 transitions to step S340. In step S340, the task execution termination operation from S300 is performed, and the measurement analysis mode M2 is terminated. Note that when it is determined in step S330 whether the total imaging time TA has been reached, if it is determined that the total imaging time TA has not been reached, execution of step S304 is started again.

When the import of the measurement results based on the sampling from the number of shots 1 to the number of shots N-1 shown in FIG. It is stored in block BN of storage device 290. When step S310 is executed, the data of the DLS analysis area is stored at the address DDN of the storage device 290. Further, the data of the SLS analysis area is stored at the address SDN of the storage device 290. In step S312, calculations for static light scattering analysis are performed based on the data stored in the address SDN. This calculation result is stored at address ARSN in step S314. Furthermore, in step S322, dynamic light scattering analysis is executed, and the calculation results are stored in address ARDN0 and address ARDN1 in step S324.

Execution by the CPU 276 moves from step S324 to step S330. In step S330, when it is determined that the end condition of the measurement analysis mode M2 shown in FIG. Execution moves to step S340. In step S340, the capture of measurement data in the measurement analysis mode M2 is finished, and the analysis operation based on the captured measurement results is finished.

In the embodiment shown in the flowchart of FIG. 16, the CPU 276 executes step S312 and step S314 before executing step S322 and step S324. This is just an example, and the order of execution of step S312 and step S314 and the execution of step S322 and step S324 may be reversed. Further, it is not necessary to always perform both steps S312 and S314 and steps S322 and S324 in succession, and they may be performed separately. Alternatively, only one of them may be performed. By executing step S304, the measurement results captured from the scattered light detection unit 200 or from the light detection camera 204, which is an example of the scattered light detection unit 200, are analyzed for both static scattered light analysis and dynamic scattered light analysis. Can be used for Therefore, by using the captured measurement results, the operator can perform both static scattered light analysis and dynamic scattered light analysis, and can also selectively perform either of them. It is possible.

The sample analysis device and analysis method based on the present invention can be used industrially.

DESCRIPTION OF SYMBOLS 11... Photodetection element, 100... Analyzer, 102... Analyzer, 110... Light source, 120... Incident light adjustment part, 122... Light amount adjustment element, 124... Polarization Element, 126... Incident light, 130... Optical path changing unit, 132... Plane mirror, 134... Plane mirror, 138... Optical aperture, 139... Aperture, 140... Light irradiation Part, 142...Condensing lens, 144...Lens adjustment mechanism, 146...Optical axis, 148...Irradiation light, 150...Sample part, 151...Incidence surface, 152... Sample cell, 153... Transmission surface, 154... Sample, 156... Analysis micro region, 157... Minimum beam spot, 158... Sample position adjustment mechanism, 160... Small angle scattered light, 162 ...Maximum small angle scattered light, 164...Transmitted light, 166...Parallel transmitted light, 168...Parallel small angle scattered light, 169...Selected parallel small angle scattered light, 170...Parallel scattered light conversion Part, 172... Lens, 174... Support device, 176... Transmitted light shielding part, 180... Scattering angle filter, 182... Aperture, 184... Specific angle parallel small angle scattered light, 185 ... Surrounding light shielding part, 186... Arc-shaped specific angle scattered light, 187... Bridge, 188... Radius, 189... Center radius, 190... Scattered light condensing part, 192...・Lens, 194...Support mechanism, 196...Small angle scattered convergent light, 200...Scattered light detection unit, 202...Photodetector, 203...Light receiving surface, 204...For light detection Camera, 205...Wiring, 206...Camera lens, 208...Semi-transparent screen, 212...Amplification/cooling section, 220...Position adjustment mechanism, 230...Photographed image, 232... - Dark part, 234... Image part, 250... Analysis part, 260... Controller, 266... Input/output device, 270... Arithmetic processing unit, 276... CPU, 280... Time Management device, 290...Storage device.

Claims (13)

a light source that generates light for measuring the sample;
a light irradiation unit that irradiates the sample with irradiation light that forms a minimum beam spot within the sample based on the light generated by the light source;
a parallel scattered light converter that receives forward or backward small-angle scattered light generated in the analytical micro region of the sample where the minimum beam spot is formed, and generates parallel small-angle scattered light based on the forward or backward small-angle scattered light; ,
a scattered light detection unit that detects the incident parallel small angle scattered light when the parallel small angle scattered light is incident;
an analysis section that includes a storage device, reads the detection results of the scattered light detection section into the storage device, and analyzes the analysis microregion of the sample based on the detection results loaded into the storage device. ,
In the scattered light detection unit, a microscope image that is an enlarged image of the state of the sample in the analysis micro region is generated based on the parallel small-angle scattered light, and the generated microscope image is further detected and stored in the memory. stored in the device,
The analysis unit is configured to analyze a dynamic scattered light component or a static scattered light component, or the dynamic scattered light component and the Analyzing both static scattered light components,
An analytical device characterized by: The analyzer according to claim 1,
The analysis unit stores a moving image of the microscope image in the storage device by repeatedly capturing the microscope image generated in the scattered light detection unit,
The analysis unit analyzes the dynamic scattered light component, the static scattered light component, or both the dynamic scattered light component and the static scattered light component based on the stored moving image. Features: Analyzer. a light source that generates light for measuring the sample;
a light irradiation unit that irradiates the sample with irradiation light that forms a minimum beam spot within the sample based on the light generated by the light source;
a parallel scattered light converter that receives forward or backward small-angle scattered light generated in the analytical micro region of the sample where the minimum beam spot is formed, and generates parallel small-angle scattered light based on the forward or backward small-angle scattered light; ,
a scattered light detection unit that detects the incident parallel small angle scattered light when the parallel small angle scattered light is incident;
an analysis section that includes a storage device, reads the detection results of the scattered light detection section into the storage device, and analyzes the analysis microregion of the sample based on the detection results loaded into the storage device. ,
The analysis unit stores the detection result of the scattered light detection unit in the storage device,
The analysis unit further determines a dynamic scattered light component or a static scattered light component, or the dynamic scattered light component and the static scattered light component, based on the detection result of the scattered light detection unit stored in the storage device. An analysis device characterized in that the analysis device analyzes both light components and performs an analysis regarding the analysis micro region of the sample,
Further, a scattering angle filter and a scattered light condensing section are provided between the parallel scattered light conversion section and the scattered light detection section,
The scattering angle filter causes the parallel small angle scattered light from the parallel scattered light converter to generate a circle forming an arc shape at a specific angle and in a plane perpendicular to the optical axis of the parallel small angle scattered light. making the arcuate specific angle scattered light enter the scattered light detection unit via the scattered light condensing unit;
The analysis device is characterized in that the scattered light detection section detects a microscopic image showing an enlarged display of the state of the analysis microregion based on the arcuate specific angle scattered light. The analyzer according to claim 3,
The analysis unit repeatedly captures the microscope image, which is an enlarged display of the state of the sample within the analysis microregion detected by the scattered light detection unit, and stores it in the storage device. An analysis device characterized in that a moving image of the microscope image, which is an enlarged display of the state of the sample, is stored in the storage device. In the analysis device according to claim 3 or 4,
The scattering angle filter is made of a material that blocks light, and has an arc-shaped aperture whose radius is a length corresponding to the specific angle with respect to the optical axis of the parallel small-angle scattered light. An analytical device featuring: The analyzer according to any one of claims 1 to 5,
The light irradiation unit includes a condensing lens for forming the minimum beam spot on the sample,
The condensing lens of the light irradiation unit and the sample are arranged along the optical axis of incident light that enters the light irradiation unit based on the light generated by the light source,
The incident light is incident on the condenser lens along the optical axis, and the incident light is a light beam having a circular cross-sectional shape, and the diameter of the light beam of the incident light is 1.6 mm or less. can be,
The numerical aperture NA of the condensing lens or the focal length f of the condensing lens that the light irradiation section has, or the numerical aperture NA of the condensing lens and the focal length f of the condensing lens are determined by the light irradiation section. is determined based on the diameter of the light beam of the incident light that is incident on the analyzer. The analyzer according to claim 6,
The light irradiation unit includes the condenser lens for condensing the irradiation light;
a lens adjustment mechanism for moving the condensing lens along the optical axis of the incident light that enters the light irradiation section;
It is equipped with
An analysis device, wherein the lens adjustment mechanism has a distance by which the condensing lens can be moved along the optical axis by ±10 mm or less, and a minimum length by which the movement can be adjusted by 50 μm or less. The analyzer according to any one of claims 1 to 7,
Analysis characterized in that the diameter of the minimum beam spot formed inside the sample by irradiating the sample with the irradiation light from the light irradiation unit is in the range of 10 μm or more and 100 μm or less. Device. In the analysis device according to one of claims 1 to 8,
It has a sample part,
The sample section includes a sample cell for holding the sample and a sample position adjustment mechanism for adjusting the position of the sample cell,
By moving the position of the sample relative to the irradiation light with the sample position adjustment mechanism, the position of the analysis micro region of the sample where the minimum beam spot is formed is set, and the setting in the sample is A microscopic image of scattered light representing the state of the analyzed microscopic region is generated in the scattered light detection unit and detected by the scattered light detection unit,
The analysis unit generates an adjustment image for adjusting a position where the minimum beam spot of the sample cell is formed by the sample position adjustment mechanism based on the detection result of the microscope image detected by the scattered light detection unit. Outputs
The analysis unit repeatedly stores the detection result of the microscope image generated in the scattered light detection unit in the storage device based on the instruction for performing the analysis,
The analysis device is characterized in that the analysis section performs the analysis based on the detection results of the scattered light detection section that are repeatedly stored in the storage device. The analyzer according to any one of claims 1 to 9,
In order to form the minimum beam spot in the sample, the maximum incident angle θi of the irradiation light irradiated from the light irradiation unit to the sample is in the range of 0.5 degrees to 20 degrees. Analyzer. The analyzer according to any one of claims 1 to 10,
a first step in which the analysis section repeatedly captures the detection results of the scattered light detection section based on set sampling conditions;
a second step of extracting a detection result of a dynamic scattered light analysis region and a detection result of a static scattered light analysis region from the repeatedly captured detection results of the scattered light detection unit;
A step of performing a calculation for dynamic scattered light analysis based on the detection result of the dynamic scattered light analysis area and a calculation for static scattered light analysis based on the detection result of the static scattered light analysis area. 3 steps and
An analysis method characterized by having the following. The analysis method according to claim 11,
After the first step of repeatedly acquiring the detection results from the scattered light detection section based on the set sampling conditions, the analysis section performs the steps of the first step during the set imaging interval. stop execution,
The analysis unit starts executing the first step again after the set imaging interval has elapsed,
The analysis unit repeats the execution of the first step until the number of executions of the first step reaches a preset number of executions, and the analysis unit repeats the execution of the first step to obtain the detection result including the microscope image generated in the scattered light detection unit. into the storage device,
Alternatively, the execution of the first step and the subsequent stop of the execution are repeated until the duration of the execution of the first step and the subsequent stop of the execution reach a preset total imaging time. , importing the detection result including the microscope image generated in the scattered light detection unit into the storage device,
The analysis unit is configured to analyze the dynamic scattered light component or the static scattered light component, or the dynamic scattered light component and the static scattered light component, based on the detection result including the microscope image captured in the storage device. Perform analysis of both components,
An analysis method characterized by: In the analysis method according to claim 11 or 12,
The analysis unit operates in an adjustment mode and a measurement analysis mode based on instructions,
In the operation in the adjustment mode, the third step of performing the calculation for the dynamic scattered light analysis and the calculation for the static scattered light analysis based on the detection result from the scattered light detection unit is stopped. , an analysis method characterized by: