JP2013214102A - Centrifugal microscope - Google Patents

Abstract

PROBLEM TO BE SOLVED: To confirm a state of a sample in a reaction process of decomposition or synthesis with video of stable imaging quality at a high frame rate in real time and to perform fluorescence observation in a centrifugal microscope.SOLUTION: On a rotary table 4, as fluorescence observation means, a centrifuge microscope mounts: a lighting device 41 arranged opposing to a rotation axis direction of a reaction device 6 and the rotary table 4; an objective lens 8a which emits excited light from the lighting device 41 onto a sample of the reaction device 6; and a dichroic beam splitter 14 which transmits the excited light from the lighting device 41 and which reflects fluorescence reflected from the sample of the reaction device 6 and transmits the fluorescence to a CCD camera 10.

Description

  The present invention is an apparatus for separating, analyzing, or synthesizing a predetermined substance from a sample by applying a centrifugal force to the sample. In particular, the state of the sample during the separation reaction or the synthesis reaction is measured in real time. The present invention relates to a centrifugal microscope that can be visually confirmed.

  Conventionally, various centrifuges (for example, centrifugal separation) for separating or synthesizing a predetermined substance (liquid, solid, gas, mixture, etc.) from this sample by applying a centrifugal force to the sample in the reactor. Devices) and methods are known, and these centrifugal devices and methods are used to purify various industrial products and pharmaceuticals, or to remove impurities from semi-finished products and reagents.

  For example, Patent Document 1 discloses as an example a blood analyzer (chip-type reactor) that separates and purifies plasma or serum from blood on a centrifuge and mixes the plasma or serum with various reagents. ing. Patent Document 2 discloses, as an example, a microsystem (a centrifugal apparatus and a method for performing microanalysis) that mixes liquid by utilizing the fact that the flow of the liquid is moved by centripetal force generated by rotation. Patent Document 3 discloses, as an example, a rotating optical biodisk provided with a fluid circuit in order to process a blood sample for clinical diagnosis and measure, for example, the amount of blood cells (red blood cell count, white blood cell count, etc.). Has been. Such a rotating optical bio-disc is loaded into a predetermined optical reader and rotated at a predetermined rotational speed for a predetermined time. Patent Document 4 discloses, as an example, a method for removing small negatively charged organic molecules from a biological sample mixture and a rotating disk reactor (device) used in the method. Patent Document 5 discloses a powder that estimates a non-electrostatic adhesion force between a powder composed of at least one kind of fine particles and an adherent material particle on a sample substrate with a centrifugal force generated by rotation of a centrifugal separator. An adhesive force measuring apparatus is disclosed.

  Here, when performing separation work on a sample using these rotary reactors, the reactor is rotated, and at the same time, the mass transfer state and reaction state in the reactor are observed (confirmed) during the work. ) Need to be kept. In particular, when measuring the adhesive force of adhesive particles as in Patent Document 5, in addition to means for measuring the number of particles before and after centrifugation, a method capable of grasping in real time the centrifugal force when the particles peel is desired. It is. In this case, a method of observing the change of the state together with the centrifugal force is necessary for measuring the adhesive force of the adhesive particles that change its adhesiveness depending on the surrounding environment, such as a cell.

  As a centrifugal apparatus equipped with such an observation mechanism, for example, in Patent Document 6, a sample chamber provided in a rotating rotor is disposed so as to cross the optical axis of an objective lens, and light emission from a pulsed light source is performed. A centrifugal microscope is disclosed in which the observation image is not stopped regardless of the rotation of the sample chamber. In Patent Documents 7 to 9, a polarization observation optical system (polarization observation optical mechanism) having an objective lens disposed so that the sample chamber (reactor) on the rotating disk crosses the optical axis thereof, and A centrifugal microscope provided with a light source that emits laser light to an optical system for polarization observation at a timing when the sample chamber crosses the optical axis of the objective lens is disclosed as an example.

  In addition, the present applicant has already filed an application described in Patent Document 10 as a centrifuge provided with an observation mechanism. The visible centrifuge and the observation device of Patent Document 10 are visible centrifuges capable of confirming the state of a sample during a separation or synthesis reaction in real time with a stable and high-quality image at a high frame rate. A rotating disk that rotates about a rotation axis, a reactor that is disposed on the rotating disk and that contains a sample and rotates together with the rotating disk, and a microscope for observing the state of the sample in the reactor. It is possible to separate or synthesize a predetermined substance from the sample by applying a centrifugal force to the sample in the reactor, and also to observe the sample state in the reactor. An imaging device for taking a microscopic image of the sample in the reactor, and wirelessly transmitting the video of the microscopic image taken by the imaging device as a moving image in real time It is disposed and video wireless transmission device for.

JP 2004-109099 A Special table 2003-533682 gazette JP 2005-509882 Gazette Special table 2006-501805 gazette JP 2006-220472 A JP-A 63-250615 JP-A-11-109245 JP-A-11-258509 Japanese Patent Laid-Open No. 11-258511 International Publication WO2008 / 078475

  By the way, depending on the sample to be observed, the behavior of this sample can be known effectively by observing the fluorescence phenomenon that occurs when light of a specific wavelength is irradiated (excited). In this case, in order to obtain a particularly good quality fluorescence observation image, it is necessary to realize effective excitation light irradiation and optical path design (fluorescence optical system) for high-sensitivity observation. Such a fluorescence optical system requires an excitation light source for generating a specific wavelength, a splitter for separating the fluorescence from the sample from the excitation light and sending it to the imaging device. However, it is difficult to realize these in a compact and high-rigidity state with the technologies of the above-mentioned patent documents, and it is difficult to realize a centrifugal microscope having a fluorescence observation function with a mechanism in which a microscope is mounted on a rotating disk. there were.

  In addition, in the centrifugal microscopes (centrifugal devices) disclosed in Patent Documents 6 to 9, a polarization observation optical mechanism is disposed outside the rotating disk, and the sample is emitted by a light source that emits light in synchronization with the rotation of the rotating disk. When observing the state, only the frame corresponding to the rotation of the rotating disk can be observed. In other words, in the case of a centrifugal microscope (centrifuge) configured so that the rotating disk rotates once per second, there is a problem that observation of one frame per second is the limit. In addition, if a microscope is placed on a rotating disk, centrifugal force is applied to the microscope's lens barrel, which increases the vibration of the lens barrel, making it difficult to perform stable observation, and for taking images. Since the same centrifugal force as that of the reactor is applied to this camera, there has been a problem that the operation of the camera becomes unstable. In particular, in the case of fluorescent observation of a sample with weak observation light, it is necessary to slow down the shutter speed to ensure image brightness in order to improve the image contrast (signal ratio S / N). However, since the sample always moves with respect to the lens, there is a problem that blurring of the image becomes large when the shutter speed is slowed down.

  The present invention has been made to solve such problems, and its purpose is to confirm the state of the sample in the reaction process of separation or synthesis in real time with a stable image quality image at a high frame rate (number of frames). An object of the present invention is to provide a centrifugal microscope capable of performing fluorescence observation.

  In order to achieve such an object, a centrifugal microscope according to the present invention includes a rotatable turntable, a reactor disposed on the outer peripheral side of the turntable and containing a sample, and a central portion of the turntable. And an imaging device capable of imaging the sample state of the reactor by rotating together with the turntable, and provided with a fluorescence observation means capable of observing fluorescence of the sample of the reactor. The observation means includes a light source using a semiconductor laser disposed on the rotating plate, an objective lens that irradiates the sample of the reactor with excitation light from the light source, and transmits the excitation light from the light source, A spectroscope that splits the fluorescence generated from the sample of the reactor with the excitation light and refracts it to send it to the imaging device, and the reflection by the spectroscope between the spectroscope and the imaging device Excitation light removing filter for removing excitation light is disposed from, it is characterized in.

  In the centrifugal microscope according to the present invention, the reactor is supported on the turntable, the light source is disposed above the turntable, and excitation light can be incident on the sample of the reactor from the light source. It is characterized by being.

  In the centrifugal microscope according to the present invention, the reactor is supported above the rotating disk, the light source is disposed below the rotating disk, and the excitation light is emitted from the light source toward the sample of the reactor. It is characterized by being possible.

  In the centrifugal microscope according to the present invention, the light source, the spectroscope, the objective lens, and the reactor are arranged in series along the rotation axis direction of the rotating disk, and the spectroscope and the excitation light removal filter And the imaging device are arranged in series along the radial direction of the rotating disk.

  The centrifugal microscope according to the present invention is characterized in that an aperture system and a condensing optical system are disposed between the light source and the spectroscope.

  In the centrifugal microscope according to the present invention, a condensing optical system is disposed between the light source and the objective lens, and the condensing optical system includes a plurality of condensing lenses disposed in series in the irradiation direction of the excitation light. It is characterized by having.

  In the centrifugal microscope according to the present invention, one or more excitations that remove excitation light in a range that enters the fluorescence wavelength region from excitation light emitted from the light source between the light source and the spectroscope. A light removal filter is provided.

  In the centrifugal microscope according to the present invention, a fluorescence observation fluid is filled between the objective lens and the reactor, and a cap for preventing leakage and scattering of the fluorescence observation fluid is disposed. It is a feature.

  In the centrifugal microscope according to the present invention, the objective lens has a numerical aperture in the range of 0.3 to 1.3 and a working distance in the range of 0.2 to 18.0 mm. .

  According to the centrifugal microscope of the present invention, the reactor for accommodating the sample is disposed on the outer peripheral side of the rotating disk, while the imaging device capable of imaging the sample state of the reactor is disposed in the central part of the rotating disk, A fluorescence observation means capable of fluorescence observation of the sample of the vessel is provided, and as the fluorescence observation means, a light source using a semiconductor laser disposed on the rotary disk facing the reactor and the rotation axis direction of the rotary disk, and excitation from the light source The objective lens that irradiates the sample of the reactor and the excitation light from the light source are transmitted, while the fluorescence generated from the sample of the reactor is split with the excitation light and refracted to reflect the fluorescence and send it to the imaging device A spectroscope is provided, and an excitation light removal filter that removes excitation light from reflected light from the spectroscope is disposed between the spectroscope and the imaging device. Accordingly, the state of the sample in the reaction process of separation or synthesis can be confirmed in real time with a stable image quality at a high frame rate (frame number), and fluorescence observation can be performed with a bright image. In addition, when excitation light from the light source passes through the spectrometer and is irradiated onto the reactor sample by the objective lens, fluorescence reflected from the reactor sample is generated, and this fluorescence is reflected by the spectrometer and the imaging device. Therefore, it is possible to perform fluorescence observation with high accuracy while making the apparatus compact. Further, even if excitation light is mixed with the fluorescence reflected by the spectroscope, this excitation light is removed by the excitation light removal filter and sent to the imaging device, and a highly accurate fluorescence image can be obtained.

  According to the centrifugal microscope according to the present invention, the reactor is supported on the rotating disk, the light source is disposed above the rotating disk, and the excitation light can be incident on the sample of the reactor from the light source. By disposing all of them on the turntable, the apparatus can be made compact.

  According to the centrifugal microscope according to the present invention, the reactor is supported above the rotating disk, the light source is disposed below the rotating disk, and the excitation light can be emitted from the light source toward the sample of the reactor. By arranging the light source below the turntable while arranging other devices on the turntable, it is possible to facilitate access to the sample and shorten the working distance between the objective lens and the sample. In addition, since the aperture ratio of the objective lens can be improved, bright observation can be achieved and the apparatus can be made compact.

  According to the centrifugal microscope according to the present invention, the light source, the spectroscope, the objective lens, and the reactor are arranged in series along the rotation axis direction of the rotating disk, and the spectroscope, the excitation light removing filter, and the imaging device are arranged on the rotating disk. Since the light source, spectroscope, objective lens, reactor, excitation light removal filter, and imaging device are efficiently arranged on the rotating disk, the device is compact and highly rigid. Can be made possible.

  According to the centrifugal microscope of the present invention, the diaphragm system and the condensing optical system are disposed between the light source and the spectroscope, so that the excitation light from the light source is constricted by the diaphragm system, and the objective is collected by the condensing optical system. Focusing is performed near the back focus of the lens, so that uniform excitation light irradiation can be achieved on the sample surface, and a compact optical path design can be achieved, and uniform and high-intensity excitation light can be achieved. Can be secured.

  According to the centrifugal microscope of the present invention, a condensing optical system is disposed between the light source and the objective lens, and a plurality of condensing lenses arranged in series in the irradiation direction of the excitation light as the condensing optical system. Therefore, the apparatus can be made compact by shortening the distance from the light source to the focal length by using a plurality of condensing lenses.

  According to the centrifugal microscope of the present invention, one or more excitation light removal filters for removing excitation light in a range that enters the fluorescence wavelength region from excitation light are disposed between the light source and the spectroscope. Therefore, the excitation light irradiated from the light source is irradiated to the sample of the reactor by removing the range in the fluorescence wavelength region by a plurality of removal filters, and a highly accurate fluorescent image can be obtained. .

  According to the centrifugal microscope according to the present invention, the fluorescence observation fluid is filled between the objective lens and the reactor, and the cap for preventing leakage and scattering of the fluorescence observation fluid is disposed. A highly accurate fluorescent image can be obtained with the fluid, and the cap can prevent leakage and scattering of the fluorescence observation fluid, thereby improving the reliability of the apparatus.

  According to the centrifugal microscope according to the present invention, the numerical aperture of the objective lens is set in the range of 0.3 to 1.3 and the working distance is set in the range of 0.2 to 18.0 mm. It is possible to achieve both high resolution of the fluorescent image and high operability of the apparatus.

FIG. 1 is a perspective view showing an example of the overall configuration of a centrifugal microscope according to the first embodiment of the present invention. FIG. 2 is a partial configuration diagram for explaining an internal configuration of a fixed component in the centrifugal microscope according to the first embodiment. FIG. 3 is a top view of the entire configuration example of the centrifugal microscope according to the first embodiment viewed from above. FIG. 4 is a longitudinal sectional view of the centrifugal microscope according to the first embodiment. FIG. 5 is an overall perspective view of the centrifugal microscope in a state where the spindle unit is mounted on the centrifugal microscope according to the first embodiment. FIG. 6 is a longitudinal sectional view of a sample mounting portion in the centrifugal microscope according to the first embodiment. FIG. 7 is a plan view schematically showing a flow path pattern of the flow path forming chip in the centrifugal microscope according to the first embodiment. FIG. 8 is a longitudinal sectional view of a diaphragm system in the centrifugal microscope according to the first embodiment. FIG. 9 is a longitudinal sectional view of a centrifugal microscope according to the second embodiment of the present invention. FIG. 10 is a perspective view showing an example of the overall configuration of a centrifugal microscope according to the third embodiment of the present invention. FIG. 11 is a longitudinal sectional view of a centrifugal microscope according to the third embodiment. FIG. 12 is a schematic diagram illustrating an optical path of the centrifugal microscope according to the third embodiment. FIG. 13 is a schematic diagram illustrating an optical path of a centrifugal microscope according to the fourth embodiment of the present invention. FIG. 14 is a longitudinal sectional view of an essential part of a centrifugal microscope according to the fifth embodiment of the present invention. FIG. 15A is a graph illustrating an absorption wavelength region by the excitation light removal filter in the centrifugal microscope according to the fifth embodiment. FIG. 15-2 is a graph illustrating an absorption wavelength region by the excitation light removal filter in the centrifugal microscope according to the fifth embodiment. FIG. 16 is a graph showing the numerical aperture and working distance of the condensing optical system in the centrifugal microscope according to the sixth embodiment of the present invention. FIG. 17 is a longitudinal sectional view of an essential part of a centrifugal microscope according to the seventh embodiment of the present invention. FIG. 18A is a longitudinal sectional view of an essential part showing a modification of the centrifugal microscope according to the seventh embodiment. FIG. 18-2 is a longitudinal sectional view of an essential part showing a modification of the centrifugal microscope according to the seventh embodiment.

  Exemplary embodiments of a centrifugal microscope according to the present invention will be described below in detail with reference to the accompanying drawings. In addition, this invention is not limited by this embodiment.

[First Embodiment]
FIG. 1 is a perspective view showing an example of the overall configuration of a centrifugal microscope according to the first embodiment of the present invention, and FIG. 2 is a diagram for explaining the internal configuration of a fixed component in the centrifugal microscope of the first embodiment. FIG. 3 is a top view of the entire configuration example of the centrifugal microscope according to the first embodiment, and FIG. 4 is a longitudinal sectional view of the centrifugal microscope according to the first embodiment. FIG. 6 is an overall perspective view of the centrifugal microscope in a state where the spindle unit is mounted on the centrifugal microscope of the first embodiment, and FIG. 6 is a longitudinal sectional view of a sample mounting portion in the centrifugal microscope of the first embodiment. 7 is a plan view schematically showing a flow path pattern of a flow path forming chip in the centrifugal microscope of the first embodiment, and FIG. 8 is a longitudinal sectional view of a diaphragm system in the centrifugal microscope of the first embodiment. is there.

  The centrifugal microscope according to the first embodiment allows a device that separates or synthesizes a predetermined substance from this sample by applying a centrifugal force to the sample, and indicates the state of the sample during the separation reaction or the synthesis reaction. It can be confirmed visually in real time.

  As shown in FIGS. 1 to 5, the centrifugal microscope A according to the first embodiment includes a rotating disk 4 that rotates around a predetermined rotating shaft 2, and a sample disposed on the outer periphery of the rotating disk 4. And a microscope 6 for visually observing the state of the sample in the reactor 6. The centrifugal microscope A then applies a centrifugal force to the sample in the reactor 6 to separate or synthesize a predetermined substance (liquid, solid and gas, or a mixture thereof) from the sample. I am letting.

  The size and shape of the centrifugal microscope A, specifically, the size and shape of the rotating disk 4 may be arbitrarily set according to the nature or number of samples to be centrifuged or centrifuged. In the embodiment, a case where the rotating disk 4 is configured as a disk having a diameter of 220 mm is assumed as an example. The reactor 6 accommodates a predetermined sample therein, and the form thereof is not particularly limited as long as the sample can be subjected to a centrifugal separation reaction or a centrifugal synthesis reaction, and the reactor 6 having an arbitrary form. However, in this embodiment, the case where the chip-type reactor 6 is applied is assumed as an example. The reactor 6 is fixed with respect to the rotating disk 4 and rotates together with the rotating disk 4 with the sample contained therein.

  In the above-described centrifugal microscope A, the microscope 8 is fixed to a predetermined position (outer peripheral portion) of the rotating disk 4 so that the state of the sample in the reactor 6 can be observed. The rotating disk 4 includes an imaging device 10 for capturing a sample microscope image in the reactor 6 captured by the microscope 8 and a captured image of the microscope image captured by the imaging device 10 as a moving image in real time. A video wireless transmission device 12 for wireless transmission is attached. In the present embodiment, as an example, the microscope 8 has an objective lens 8a that captures the state of the sample in the reactor 6, and an optical path for transmitting a microscope image captured by the objective lens 8a to the imaging device 10. And a lens barrel 8b formed in the above. In addition, various imaging devices capable of capturing a microscopic image of the sample state in the reactor 6 captured by the microscope 8 can be applied to the imaging device 10, but in this embodiment, imaging is performed. A case where a CCD camera is applied as the device 10 is assumed as an example.

  In this case, in the microscope 8, the optical path of the microscope 8 is partially refracted at a predetermined angle with respect to the disk surface (the upper surface in FIG. 4) 4 a of the rotating disk 4 in the lens barrel 8 b, and the imaging device ( A CCD camera (hereinafter referred to as a CCD camera) 10 is positioned in the vicinity of the rotation center of the turntable 4 so that the above-described microscope image can be taken on the optical path of the microscope 8 refracted at a predetermined angle. In the following description, the optical path of the microscope 8 described above is referred to as an observation optical path, and the light traveling along the observation optical path is referred to as observation light.

  In the present embodiment, as an example, as shown in FIG. 4, the mirror 14 is tilted with respect to the observation optical path by a predetermined angle arbitrarily set according to the refraction angle of the observation optical path into the lens barrel 8b of the microscope 8. Are arranged. In the configuration shown in FIG. 4, in the centrifugal microscope A, the objective lens 8a of the microscope 8 captures the sample state in the reactor 6 from above in the vertical direction (the vertical direction in the figure), and the center of rotation of the turntable 4 In this vicinity, a microscope image captured by the objective lens 8a is captured by the CCD camera 10 from a direction (horizontal direction) parallel to the disk surface 4a of the rotating disk 4. For this reason, when the mirror 14 is tilted backward at an angle of about 135 ° with respect to the entrance direction of the observation light n in FIG. 4 and disposed in the lens barrel 8b of the microscope 8, the entered observation light n is about 90%. The light is refracted by only .degree. The microscope 8 may have a configuration in which an observation optical path refracted substantially at right angles is formed in the lens barrel 8b by refracting the lens barrel 8b at substantially right angles.

  In this case, by positioning the microscope 8 at the peripheral edge of the rotating disk 4, the observation light n entering from the vertical direction is parallel to the disk surface 4 a of the rotating disk 4 from the circumferential direction of the rotating disk 4 toward the center by the mirror 14. The traveling direction can be changed so as to be refracted. As a result, the microscope 8 has a structure in which the observation light n (that is, a microscope image) reaches (converges) toward the central portion of the rotating disk 4, that is, the direction of the rotation center of the rotating disk 4. Is positioned at the arrival (convergence) destination of the observation optical path, so that the CCD camera 10 captures the observation light n of the microscope in the vicinity of the rotation center of the turntable 4, specifically, a microscope image can be taken. It can be configured.

  For this reason, the CCD camera 10 can be positioned in the vicinity of the rotation center of the turntable 4, and even if a centrifugal force is generated by the rotation of the turntable 4, the centrifugal force acting on the CCD camera 10. Therefore, it is possible to always stably shoot a microscopic image in the CCD camera 10 without the performance of the CCD camera 10 being hindered by the centrifugal force and the microscopic image at the time of photographing is not blurred. Become.

  In addition, as described above, the microscope 8 has a structure in which the observation optical path is refracted by the mirror 14, whereby the height of the lens barrel 8 b of the microscope 8 (the vertical distance in FIG. 4) can be suppressed. Thereby, even if the rotation vibration is generated by the rotation of the turntable 4, the rigidity of the microscope 8 with respect to the rotation vibration can be increased, and the sample state in the reactor 6 which is always stable in the microscope 8 can be observed. Can be performed. However, in order to suppress the height of the lens barrel 8b, it is preferable that the microscope 8 has a structure in which the refraction angle of the observation optical path is larger than 0 ° and not larger than 90 °.

  The microscope 8 has a structure in which the lens barrel 8b can move up and down in the vertical direction (vertical direction-up and down direction in FIG. 4) with respect to the reactor 6, and by forming such a structure, The distance (focal length) between the reactor 6 (specifically, the sample) and the objective lens 8a can be adjusted. In this case, the rotary disk 4 is provided with a guide 20 (hereinafter referred to as a Z-axis guide) 20 extending in a vertical direction with respect to the disk surface 4a. The microscope 8 has a structure for adjusting the focal length between the sample and the objective lens 8a.

  In the present embodiment, the objective lens 8a of the microscope 8 and the reactor 6 are accommodated in the same component (hereinafter referred to as a fixed component) 16, and the accommodated objective lens 8a and the reactor 6 are fixed. Relative displacement due to external vibration between the objective lens 8a and the reactor 6 (specifically, rotational vibration generated by rotation of the rotating disk 4) is minimized by fixing the component 16 to the rotating disk 4 integrally. I am letting. Thereby, the microscope 8 can always stably capture the sample state in the reactor 6 as a clear image without blurring, and can accurately and reliably observe the state of the sample in the separation process or the synthesis process. It becomes.

  In this case, as shown in FIG. 2, a predetermined illumination device (for example, an edge-type LED (Light Emitting Diode) backlight) 22 is provided inside the fixed component 16, and the sample is placed on the objective of the microscope 8. Observation is possible in a state of being transmitted through the illumination device 22 from the side opposite to the lens 8a. Thereby, the state of the sample in the reactor 6 can be observed more clearly, and the sample state can be captured as a clearer microscope image by the objective lens 8a. Note that the LED 22a, which is the light source of the illumination device (edge-type LED backlight) 22, as in the CCD camera 10 described above, rotates the rotating disk 4 in order to reduce the action of centrifugal force generated by the rotation of the rotating disk 4. It is positioned near the center.

  Here, the specific configuration of the illumination device 22 is not particularly limited as long as the illumination device 22 can observe the sample while illuminating and transmitting the sample. For example, an arbitrary illumination device may be selected according to the nature or type of the sample, and as an example, in this embodiment, LED illumination (edge type backlight) MEBL-CW25 manufactured by Moritex Corporation is used. ing. However, for example, a lighting device having performance equivalent to or higher than that of the lighting device 22 may be used.

  The fixed component 16 adjusts the focal length between the objective lens 8a of the microscope 8 and the sample, and is set to an appropriate distance, and the relative position between the objective lens 8a and the sample, specifically, the objective lens 8a. The height of the sample is fixed. Thereby, during the separation reaction or the synthesis reaction, the height of the objective lens 8a of the microscope 8 with respect to the sample can be kept constant, and the state of the sample can be observed stably.

  Here, in the above-described centrifugal microscope A, it is necessary to further improve the magnification and resolution in order to make it possible to observe a finer object than a specific substance such as a micro organism. Therefore, in this embodiment, in order to improve the resolution more easily, the sample mounting portion of the centrifugal microscope A is configured as described below.

  As shown in FIG. 6, the sample mounting portion 60 includes a sample table 61 having a pinhole 61 a at the substantially center, a glass plate, a resin plate, and the like disposed on the surface 61 b of the sample table 61 so as to cover the pinhole 61 a. And is provided between the reactor 6 as the observation object and the backlight type illumination device 22. The reactor 6 is placed and mounted on the cover member 62.

  The sample stage 61 is configured such that the back surface on the illumination device 22 side with respect to the flat surface 61b is in a mortar shape with the pinhole 61a as the center, that is, a substantially conical concave surface 61c inclined toward the pinhole 61a. The outer peripheral portion of the substantially conical concave surface 61 c is placed on the illumination device 22. The sample stage 61 can be composed of a light metal material such as aluminum or a resin material. The substantially conical concave surface 61c inside the sample stage 61 is black-processed, for example.

  According to this sample mounting part 60, by installing the sample stage 61 provided with the pinhole 61a on the illumination device 22, a part of the backlight illumination light P from the illumination device 22 is transmitted through the pinhole 61a. The transmitted light is irradiated to the observation region 6a of the observation object (reactor 6) from the back side, and the light transmitted through the observation region 6a enters the objective lens 8a as the observation light n, and the sample in the reactor 6 is I can observe. In this way, almost all of the transmitted light that has passed through the pinhole 61a is applied only to the observation region 6a, so that the contrast of the image is improved and an image with high resolution can be obtained.

  In the present embodiment, the sample stage 61 is provided with a pinhole 61a, and the transmitted light transmitted through the pinhole 61a of the backlight illumination light P is irradiated only to the observation region 6a, thereby improving the contrast and The resolution can be improved. In addition, since the substantially conical concave surface 61c inside the sample stage 61 is configured in a mortar shape with the pinhole 61a as the center, the illumination light from the illuminating device 22 can be efficiently introduced into the pinhole 61a and the substantially conical shape. Generation of scattered light is prevented by the black treatment of the concave surface 61c.

  In addition, the above-mentioned centrifugal microscope A is a reactor 6, for example, as shown in FIG. 7, a dumbbell-shaped fine flow path pattern (circular solution tanks are provided at both ends, and the solution tanks are connected by a straight flow path. A flow channel device in which a flow channel having a shape is formed, and the flow channel device is installed. This channel device has a structure in which a PDMS resin on which a fine channel pattern is formed is attached on a glass substrate. The channel pattern is, for example, a shape in which circular solution layers having a diameter of 3 mm are connected by a straight channel having a width of 700 μm, and the depth is about 120 μm. On one side, holes for introducing a solution are formed.

  The channel device is attached to the centrifugal microscope A so that the straight channel portion can be observed with the CCD camera 10 with the solution introduction hole on the anti-centrifugal side. At this time, the flow path is filled with the solution. When a solution is introduced from the solution introduction hole and the centrifugal microscope A is driven, a specific substance moves to the centrifugal solution tank through a straight channel by centrifugal force. In the centrifugal microscope A, the moving speed v of the specific substance can be accurately measured because the speed at which the specific substance moves in the straight flow path can be measured at an arbitrary number of rotations (centrifugal force).

  By the way, in this Embodiment, the centrifugal microscope A has the fluorescence observation means which can carry out the fluorescence observation of the sample of the reactor 6. FIG. That is, by irradiating (exciting) light of a specific wavelength to a sample and observing the fluorescence phenomenon that occurs at this time, the behavior of this sample can be known effectively. In this case, in order to obtain a particularly good quality fluorescence observation image, it is necessary to realize effective excitation light irradiation and optical path design (fluorescence optical system) for high-sensitivity observation. Such a fluorescence optical system requires an excitation light source for generating a specific wavelength, a splitter for separating the fluorescence from the sample from the excitation light and sending it to the imaging device.

  In the centrifugal microscope A of the present embodiment, an epi-illumination device that illuminates the sample from the reactor 6 from above is added. That is, in the microscope 8, a predetermined illumination device (for example, a bullet-type LED (Light Emitting Diode) 41) is provided on the upper part of the lens barrel 8b so that the sample can be observed in an illuminated state from above. I have to. In addition, the lens barrel 8b is located between the illumination device 41 and the mirror 14, and a diaphragm system 42 and a condensing optical system 43 are arranged in series from above.

  As shown in FIG. 8, the diaphragm system 42 includes a plate 44 having a pinhole 44a at the center thereof, and a glass plate, a resin plate, or the like disposed on the surface (lower surface) 44b of the plate 44 so as to cover the pinhole 44a. The cover member 45 which consists of is provided. The cover member 45 can be omitted. The plate 44 is configured such that the back surface (upper surface) on the illuminating device 41 side is a mortar shape with the pinhole 44a as the center, that is, a substantially conical concave surface 44c inclined toward the pinhole 44a. The lighting device 41 is disposed so as to face the substantially conical concave surface 44c. The plate 44 can be made of a light metal material such as aluminum or a resin material, and the inner substantially conical concave surface 44c is black-treated, for example. The diaphragm system 42 having the pinhole 44a may be set according to the type of the illumination device 41, and the diaphragm system 42 may be omitted when securing a large amount of light.

  The condensing optical system 43 condenses the illuminating light of the illuminating device 41 that has passed through the pinhole 44 a of the aperture system 42 and transmits it to the mirror 14. The upper mirror 14 of the objective lens 8a is a dichroic beam splitter that functions as a spectroscope. The illumination light m from the illumination device 41 passes through the dichroic beam splitter 14, and the sample in the reactor 6 passes through the objective lens 8a. Irradiate. Reflected light (observation light) n from the sample is reflected by the dichroic beam splitter 14 through the objective lens 8 a and reaches the CCD camera 10.

  Here, the specific configuration of the illumination device 41 is not particularly limited as long as the illumination device 41 can be observed in a state where the sample is illuminated. For example, an arbitrary illumination device may be selected according to the nature and type of the sample.

  In this embodiment, since a fluorescent substance (fluorescent reagent) such as FITC (fluorescein isothiocyanate) or GFP (green fluorescent protein) generally known in biological experiments is used as a target, a blue LED (wavelength) is used as a light source. : 460 nm to 490 nm). In this optical system, the sample is irradiated with light of this wavelength as excitation light, and fluorescence of 510 nm to 550 nm generated by FITC or GFP is observed. As LED illumination that can be used as this excitation light source, for example, high power spot illumination MCEP-CB8 manufactured by Moritex Co., Ltd., coaxial / spot illumination IHV-27BmkII manufactured by IMAX Co., Ltd. can be used. It is preferable to be as compact and lightweight as possible. Moreover, it is good also as what attached the cover to high-intensity bullet-type LED instead of the above-mentioned commercial lighting apparatus. In addition, it is preferable that the light source has been collimated with a lens or the like in advance, and a semiconductor laser or the like can also be suitably used.

  The condensing lens used by the condensing optical system 43 is, for example, an TECH SPEC φ12 plano-convex lens (PCX) or a biconvex lens (DCX) (focal length 12 to 100 mm) manufactured by Edmund Optics. Yes. The objective lens 8a of the microscope 8 uses, for example, SLMPL 50 (numerical aperture: 0.45, working distance: 15.0 mm, mass: 91 g) manufactured by Olympus Corporation. It is desirable to select a large number of lenses. The dichroic beam splitter 14 uses Semlock FF505-SDi01.

  As described above, in the centrifugal microscope A of the present embodiment, the optical system uses the illumination device 41, the aperture system 42, the condensing optical system 43, the dichroic beam splitter 14, and the objective lens 8a, thereby realizing Keller illumination. ing. The Keller illumination enables uniform and strong illumination with a large aperture ratio (numerical aperture), and adjusts the aperture system 42 and the condensing optical system 43 so that the focal position on the rear surface (light source side) of the objective lens 8a. An image of the light source is formed. Since the aperture stop is placed at the position of the focal point on the light source side of the objective lens 8a of the sample, the imaged light passes through the objective lens 8a and is uniformly irradiated on the sample surface. The working distance between the objective lens 8a and the sample is adjusted so that the image of the field stop is formed on the sample. For this reason, the aerial image of the field stop is formed in the sample, but the brightness in the sample is also uniform because the brightness is more uniform at the position of the field stop.

  Therefore, the excitation light Q from the illumination device 41 is narrowed by passing through a pinhole 44a (for example, about 0.2 mm in diameter) of the diaphragm system 42, and the diaphragm light is condensed by the condensing optical system 43. The light enters the objective lens 8a. In this case, the aperture light is adjusted by the condensing optical system 43 so that it passes through the dichroic beam splitter 14 and is condensed near the back focus (rear focal plane) of the objective lens 8a. At this time, an aperture stop may be provided at the back focus position, and if a field stop is provided before and after the aperture stop, the illumination system can be easily adjusted, and image deterioration due to stray light or the like can be suppressed. When the sample is irradiated with the excitation light by the objective lens 8a, the reflected light (fluorescence) generated from the specific substance returns to the dichroic beam splitter 14, and has a wavelength of 505 nm or more derived from the fluorescence emission of the specific substance. Only light is reflected so as to be refracted and sent to the CCD camera 10 which is an imaging device.

Specifically, when excitation light (wavelength λ ₁ : 460 nm to 490 nm) of the blue LED is irradiated from the illumination device 41 to the sample in the reactor 6, it is generated as a part of the reflected light by the FITC or GFP of the sample. Fluorescence (wavelength λ ₂ : 510 nm to 550 nm) is obtained. The excitation light (wavelength λ ₁ ) and fluorescence (wavelength λ ₂ ) reflected by the sample are dispersed by the dichroic beam splitter 14, and only the fluorescence (wavelength λ ₂ ) is refracted and sent to the CCD camera 10.

  In the present embodiment, a camera image (captured image) of the microscope image captured by the CCD camera 10 is wirelessly transmitted to the rotating disk 4 in real time as well as the CCD camera 10 that captures the above-described microscope image as a movie. A video wireless transmission device 12 is attached. In addition, an excitation light removal filter 46 that removes excitation light included in the fluorescent light from the reflected light from the dichroic beam splitter 14 is disposed between the dichroic beam splitter 14 and the CCD camera 10.

The excitation light removal filter 46 removes the excitation light component contained in the reflected light refracted by the dichroic beam splitter 14. That is, the excitation light removal filter 46 is an absorption filter that transmits only light of 500 nm or more of the reflected light, and transmits the fluorescence (wavelength λ ₂ ) and absorbs the excitation light (wavelength λ ₁ ). Semlock FF01-512 / 630-25 is used.

  As described above, by adopting a wireless system instead of a wired system as a system for transmitting an image captured by the CCD camera 10 to an external receiver (not shown), the CCD camera 10 and the video wireless system together with the rotating disk 4 are used. Even when the transmission device 12 is rotated, there is no need to take out signal lines directly from them, and there is no need to consider handling of signal lines. As a result, the peripheral structure of the CCD camera 10 and the video wireless transmission device 12 can be easily simplified.

  In addition, since it is not necessary to consider the handling of the signal line, the CCD camera 10 and the video wireless transmission device 12 are configured to rotate together with the rotating disk 4 (specifically, the sample in the microscope 8 and the reactor 6). The microscope image can be taken by the CCD camera 10 at an arbitrary high frame rate (frame number) without being restricted by the rotation speed of the turntable 4, and the camera image of the taken microscope image is received externally. It can be transmitted to a device (not shown).

  Accordingly, by providing a display device such as a liquid crystal panel or a CRT (Cathode Ray Tube) display as the external receiving device, the above-described camera image (that is, the state of the sample inside the reactor 6) is applied. The sample separation reaction or synthesis reaction can be advanced while confirming in real time on the display. In addition, by analyzing the camera video recorded via a personal computer, the behavior of the sample (specifically, its internal materials, separated materials, synthetic materials, etc.) is monitored, and the optimal rotation conditions of the turntable 4 are monitored. (Another way of understanding is to estimate the optimum magnitude of the centrifugal force acting on the sample) and to control the rotation of the turntable 4 under the estimated optimum conditions (for example, rotation speed and rotation time). It is also possible.

  In this case, the video wireless transmission device 12 is positioned in the vicinity of the rotation center of the turntable 4 in order to reduce the action of the centrifugal force generated by the rotation of the turntable 4, as with the CCD camera 10 described above. Video data of a camera image taken by the camera 10 is wirelessly transmitted from a predetermined antenna 18 to an external receiver (a display device such as a liquid crystal panel or a CRT display connected to the receiver). Similarly, the antenna 18 is also configured to rise in the vicinity of the center of rotation of the rotating disk 4, specifically, on an extension line of the rotating center of the rotating disk 4 in order to reduce the action of centrifugal force generated by the rotation of the rotating disk 4. Yes.

  When the video wireless transmission device 12 wirelessly transmits the video data (video signal) of the camera video to an external receiver (not shown), the video data transmission speed (bit rate) and data format (frequency, compression / non-compression) Or the like) may be arbitrarily set according to the usage mode and usage conditions of the centrifugal microscope A. For example, in the present embodiment, as an example, a camera image of a microscope image taken by the CCD camera 10 is converted into a video data of an uncompressed digital signal having a frequency of 2.4 GHz by the video wireless transmission device 12, and the video Data is transmitted wirelessly to an external receiver. As a result, the video signal transmitted from the video wireless transmission device can be transmitted to the external receiving device without losing the video signal, and the external receiving device can perform separation while confirming the sample state with a clear and stable video. The reaction or the synthesis reaction can proceed. However, the video data transmitted from the video wireless transmission device 12 to the external reception device may be a compressed signal or an analog signal instead of the above-described uncompressed digital signal.

  Here, the CCD camera 10 is not particularly limited in its specific configuration as long as it can take a microscopic image of the sample state in the reactor 6 captured by the microscope 8. For example, an arbitrary CCD camera may be selected according to the usage mode or usage conditions of the centrifugal microscope A. As an example, in the present embodiment, the color board camera MSC-90 manufactured by MOSWELL Co., Ltd. (minimum subject) Illuminance / 0.5 lx.) Is used. Further, as a high-sensitivity camera for fluorescence observation, WATEC WAT902HB2S (minimum subject illumination / 0.0003 lx.) May be used. However, for example, a CCD camera having performance equivalent to or higher than that of the CCD camera 10 may be used.

  Further, the specific configuration of the video wireless transmission device 12 is not particularly limited as long as it can wirelessly transmit a camera image of a microscope image taken by the CCD camera 10 to an external receiving device (not shown). . For example, any video wireless transmission device may be selected according to the usage mode or usage conditions of the centrifugal microscope A, and as an example, TRX24mini manufactured by Aiden Videotronics Co., Ltd. is used in the present embodiment. . However, for example, a video wireless transmission device having performance equivalent to or higher than that of the video wireless transmission device 12 may be used.

  In addition, as shown in FIG.1 and FIG.3, various electrical components provided in the centrifugal microscope A such as the CCD camera 10, the wireless video transmission device 12, and the illumination device 22 described above are predetermined power supply devices (for example, Battery) 24. In this case, the power supply device 24 generates power that can normally operate such various electrical components (such as the CCD camera 10, the video wireless transmission device 12, and the illumination device 22) during the separation reaction or synthesis of the sample. The specific configuration is not particularly limited as long as it can be stably supplied during the reaction. For example, an arbitrary power supply device may be selected according to the magnitude of power required for the various electrical components described above. As an example, in this embodiment, UBBP01 (a battery made by ULTRA LIFE Co., Ltd.) A voltage of 3.7 V and a battery capacity of 1.8 Ah) are used. However, for example, a power supply device having performance equivalent to or higher than that of the power supply device 24 may be used.

  In the present embodiment, four such power supply devices (batteries) 24 are used in series, and these four batteries 24 are moved to positions symmetrical with respect to the rotation center of the turntable 4 (with a phase difference of 180 °). 2) Evenly arranged two by two and arranged with a phase difference of 90 ° with respect to the microscope 8, the reactor 6 and the fixed part 16 (see FIG. 3). In this case, as an example, the battery 24 is embedded in a mounting portion formed by recessing the disk surface 4 a of the rotating disk 4, fixed by the plate-like member 26, and attached to the rotating disk 4.

  Here, in the centrifugal microscope A, the rotating shaft 2 of the rotating disk 4 is rotated by a predetermined driving device (for example, a spindle motor) (not shown) and is rotatably supported by various bearings 27. FIG. 4 shows, as an example, a configuration in which the rotating shaft 2 is supported by a rolling bearing 27 in which balls are used as rolling elements. In this case, the rolling bearing 27 may be a roller bearing to which various rollers (cylindrical rollers, tapered rollers, spherical rollers, etc.) are applied as rolling elements in addition to various ball bearings in which balls are applied as rolling elements. In the configuration shown in FIG. 4, the rotary shaft 2 is supported by two bearings 27, but the rotary shaft 2 may be supported by one bearing 27 or by three or more bearings 27. You may support.

  Note that an air bearing is applied as the bearing 27 in place of the above-described various rolling bearings, and the rotary shaft 2 is rotatably supported by the air bearing, so that the rotational vibration generated when the rotating disk 4 rotates is greatly reduced. More preferably, the rotational vibration of the microscope 8 and the CCD camera 10 can be suppressed, and stable observation and photographing of the sample state during the separation reaction or the synthesis reaction can be performed. Here, as an example, the air bearing has a structure in which the rotary shaft 2 is rotatably supported by a cylindrical housing that is positioned so as to cover the entire outer peripheral surface of the rotary shaft 2. Air is blown toward the outer peripheral surface of the rotating shaft 2 from a plurality of outlets (spout holes) provided on the peripheral surface (the surface facing the outer peripheral surface of the rotating shaft 2), and the inner peripheral surface of the housing and the outer peripheral surface of the rotating shaft 2 Is kept in a non-contact state, the rotating shaft 2 can be rotated very smoothly. In this case, as the air spindle unit, for example, GBS100H manufactured by NSK Ltd. can be used.

  Further, in the present embodiment, the rotary shaft 2 and the bearing 27 that rotatably supports the rotary shaft 2 are configured as a spindle unit 28 that is integrated with the housing, and the spindle unit 28 is attached to the rotary disk 4. As a result, the rotating disk 4 is rotated about the rotating shaft 2. In this case, the central portion of the turntable 4 is convex with a predetermined size toward the upper side (the side on which the microscope 8, the reactor 6, the CCD camera 10, the video wireless transmission device 12, etc. are disposed). A spindle unit mounting portion 4b that protrudes and is formed by recessing the lower side of the rotating disk 4 (the side opposite to the side on which the above-described components are disposed) into the convex shape is provided. 28 is inserted into the spindle unit mounting portion 4 b from the lower side of the rotating disk 4 and attached to the rotating disk 4.

  Thus, the microscope 8, the reactor 6, the CCD camera 10, the video wireless transmission device 12, and the like are arranged by making the centrifugal microscope A cover the spindle unit 28 with the turntable 4. It is possible to reduce the distance between the rotational center of gravity during rotation of the turntable 4 and the shaft support portion where the rotary shaft 2 is rotatably supported by the bearing 27, and to effectively reduce the rotational moment generated in the shaft support portion. Can do.

  In addition, in this Embodiment mentioned above, although it did not mention in particular about the material of the structural member of the centrifugal microscope A, it selects and uses various raw materials arbitrarily according to the use aspect, usage purpose, etc. of the centrifugal microscope A. do it. As an example, in the present embodiment, various attachment members for attaching the material of the turntable 4 and the microscope 8, the reactor 6, the CCD camera 10, and the video wireless transmission device 12 to the turntable 4 are provided. By using high-strength aluminum alloy (A2017), the weight of these members is reduced while ensuring sufficient rigidity during rotation. In this case, various Ti alloys, Mg alloys, and the like may be used as the material for reducing the weight.

  Further, in the present embodiment, in order to make the weight balance of the centrifugal microscope A at the time of rotation uniform and to reduce the swinging stress on the spindle unit 28 generated at the time of rotation, the microscope 8 and the reactor 6 are A predetermined balance weight 30 is provided at a position that is substantially symmetrical with respect to the arrangement position and the rotation center (on the opposite side to the rotation center). The weight and position of the balance weight 30 are the weights of various members such as the microscope 8, the reactor 6, the CCD camera 10, and the video wireless transmission device 12 disposed on the turntable 4, and their balance (center of gravity). Accordingly, adjustment may be made so that the above-described swinging stress on the spindle unit 28 is reduced.

  Next, a description will be given of the operational effects of attaching and rotating the reactor 6 to the centrifugal microscope A as described above, and causing the generated centrifugal force to act on the fluid to drive and flow the fluid.

  The reactor 6 does not require fluid connection with an external device (when a pump is used for fluid drive) and electrical connection (for example, when electroosmotic flow is used for fluid drive), and the structure of the reactor 6 is reduced. It can be simplified. As an effect, the handling of the reactor 6 becomes easy, it is easy to automate, and the analysis speed is improved. Further, the reactor 6 can be further downsized, and analysis with a smaller sample becomes possible. In this case, since the cells cannot be crushed electrically, they are crushed by mechanical collision. In addition, since the peripheral device can be reduced in size, the entire measurement system can be reduced in size.

  Further, the fluid can be driven without being affected by the chemical characteristics of the sample. In particular, it is possible to drive (analyze) even a sample mainly composed of a solution that is easily electrolyzed by applying an electric field. Moreover, it can be used without worrying about these effects even for samples that may change due to electrical stimulation, and the application range is widened.

  In addition, the centrifugal effect of the sample can be generated at the same time, and separation by the specific gravity of the sample is possible.

  Furthermore, by using the centrifugal microscope of the present embodiment, it is possible to grasp the detection state without missing information (frame dropping) even in a reaction at a low rotation speed (low centrifugal force) region. Can do. At the same time, when observing faint light such as fluorescence observation, even if the exposure time is extended, the observation-side objective lens and the sample rotate together, so that there is little blurring of the image and high-precision observation is possible. .

  In addition, the centrifugal microscope A of the present embodiment is provided with a fluorescence observation means capable of fluorescence observation of the sample of the reactor 6, and the state of the sample in the separation or synthesis reaction process is set at a high frame rate (number of frames). In addition to being able to confirm in real time with a stable image quality image, fluorescence observation can be performed.

  And as this fluorescence observation means, the illuminating device 41 disposed on the rotating disk 4 so as to oppose the rotating axis direction of the reactor 6 and the rotating disk 4, and the excitation light from the illuminating device 41 as a sample of the reactor 6 An objective lens 8 a to be irradiated and a dichroic beam splitter 14 that transmits the excitation light from the illumination device 41 and reflects the fluorescence reflected from the sample of the reactor 6 to the CCD camera 10 are provided. Accordingly, when the excitation light from the illumination device 41 passes through the dichroic beam splitter 14 and is irradiated onto the sample of the reactor 6 by the objective lens 8a, fluorescence reflected from the sample of the reactor 6 is generated, and this fluorescence is dichroic. Reflected by the beam splitter 14 and sent to the CCD camera 10, high-accuracy fluorescence observation can be performed while making the apparatus compact.

  Further, an excitation light removing filter 46 for removing excitation light from the reflected light by the dichroic beam splitter 14 is disposed between the dichroic beam splitter 14 and the CCD camera 10, and excitation is performed on the fluorescence reflected by the dichroic beam splitter 14. Even if light is mixed, the excitation light is removed by the excitation light removal filter 46 and sent to the CCD camera 10, and a highly accurate fluorescent image can be obtained.

  The illuminating device 41, the dichroic beam splitter 14, the objective lens 8a, and the reactor 6 are arranged in series along the rotation axis direction of the rotating disk 4, and the dichroic beam splitter 14, the excitation light removing filter 46, and the CCD camera 10 are arranged. Are arranged in series along the radial direction of the rotating disk 4. The reactor 6 is supported on the turntable 4, the illumination device 41 is disposed above the turntable 4, and the excitation light is incident on the sample of the reactor 6 from the illumination device 41. For this reason, the illumination device 41, the dichroic beam splitter 14, the objective lens 8a, the reactor 6, the excitation light removal filter 46, and the CCD camera 10 are efficiently arranged on the rotating disk 4, thereby reducing the size and rigidity of the device. Can be possible. Further, by arranging all the devices on the turntable 4, it is possible to make the devices compact.

  Further, an aperture system 42 and a condensing optical system 43 are disposed between the illumination device 41 and the dichroic beam splitter 14, and excitation light from the illumination device 41 is stopped by the aperture system 42, and the condensing optical system. The light is condensed near the back focus of the objective lens 8a by 43, so that a compact optical path design can be achieved and uniform and high amount of excitation light can be secured.

[Second Embodiment]
FIG. 9 is a longitudinal sectional view of a centrifugal microscope according to the second embodiment of the present invention. In addition, the same code | symbol is attached | subjected to the member which has the same function as what was demonstrated in embodiment mentioned above, and the overlapping description is abbreviate | omitted.

  In the second embodiment, as shown in FIG. 9, the centrifugal microscope generates a relatively low centrifugal force (hypergravity about several times the gravity) for the purpose of measuring the gravitational behavior of micro-organisms. However, in this embodiment, since the centrifugal microscope B is equipped with a microscope system on a rotating disk, a high-frame-rate movie is used regardless of the number of rotations. It can be photographed and is particularly suitable as a device for observing the behavior of micro-organisms that move quickly. In the same application, in order to reproduce a sudden change in the gravitational environment, a device having a quick response to a change in the rotational speed is required. Therefore, the centrifugal microscope B may be driven directly by the direct drive motor.

  That is, the rotating part 51 located on the outer peripheral side of the direct drive motor 50 is attached to the rotating disk 4 to constitute a motor built-in type. Thereby, it is possible to improve the responsiveness of the rotational speed change and to reduce the size of the apparatus. As the direct drive motor 50, for example, a PS motor (1006 series) manufactured by NSK Ltd. can be used.

  Further, a slip ring 53 is provided on the mounting portion 4b above the fixed portion 52 inside the direct drive motor 50. For example, when the image sampling interval is long and the image is not visible, or a sample having a very slow gravity reaction is used as a target. When continuous observation of time is necessary, a video signal may be transmitted to the outside via the slip ring 53, or power may be supplied. Further, by using the slip ring 53, it is possible to carry out a centrifugal load test while suppressing the influence of radio waves on a biological sample in the case of an external device or a biological application. Furthermore, it can be used in an environment where radio waves cannot be used, for example, in medical equipment, in a space station, or in the vicinity of other radio interference equipment.

[Third Embodiment]
FIG. 10 is a perspective view showing an example of the overall configuration of a centrifugal microscope according to the third embodiment of the present invention, FIG. 11 is a longitudinal sectional view of the centrifugal microscope of the third embodiment, and FIG. It is the schematic showing the optical path of the centrifuge microscope of embodiment. In addition, the same code | symbol is attached | subjected to the member which has the same function as what was demonstrated in embodiment mentioned above, and the overlapping description is abbreviate | omitted.

  In the third embodiment, as shown in FIGS. 10 to 12, the centrifugal microscope C is an inverted centrifugal microscope, and is arranged on the outer peripheral portion of the rotating disk 4 to store the sample and to rotate the rotating disk 4. And a microscope 6 for visually observing the state of the sample in the reactor 6. Then, the centrifugal microscope C separates or synthesizes a predetermined substance (liquid, solid and gas, or a mixture thereof) from the sample by applying a centrifugal force to the sample in the reactor 6. I am letting.

  In the centrifugal microscope C, the microscope 8 is fixed to a predetermined position (outer peripheral portion) of the rotating disk 4 so that the state of the sample in the reactor 6 can be observed. The rotating disk 4 includes an imaging device (CCD camera) 10 for capturing a sample microscope image in the reactor 6 captured by the microscope 8 and a captured image of the microscope image captured by the imaging device 10. A wireless video transmission device 12 for wirelessly transmitting a moving image in real time is attached. In this case, in the microscope 8, the optical path is partially refracted at a predetermined angle with respect to the surface of the turntable 4 (the upper surface in FIG. 11) 4a, and the CCD camera 10 is refracted at a predetermined angle. It is positioned in the vicinity of the rotation center of the turntable 4 so that the above-described microscope image can be taken on the optical path 8.

  In the centrifugal microscope C of the present embodiment, the rotating disk 4 is provided with a guide (not shown) extending in a vertical direction with respect to the disk surface 4a, and the reactor 6 is mounted. The sample holder 71 is supported so as to be vertically slidable along the guide, and the focal length of the microscope 8 is adjusted by sliding the sample holder 71. The reactor 6 is attached to the lower surface of the sample holder 71, and an ascending illumination device for illuminating the sample from below is added. That is, in the microscope 8, a predetermined illumination device (for example, a bullet-type LED) 41 is provided on the lower surface of the rotating disk 4 so that the sample can be observed in an illuminated state from below.

  A lens barrel 8b of the microscope 8 is integrally fixed to the surface 4a of the rotating disk 4 facing the illumination device 41, and a dichroic beam splitter 14 functioning as a spectroscope is fixed to the lens barrel 8b. The objective lens 8 a is fixed above the dichroic beam splitter 14. In this case, the dichroic beam splitter 14 is disposed in the lens barrel 8b of the microscope 8 so as to be inclined with respect to the observation optical path by a predetermined angle arbitrarily set according to the refraction angle of the observation optical path. Although not shown, the lens barrel 8b or the turntable 4 is located between the illumination device 41 and the dichroic beam splitter 14, and a diaphragm system 42 and a condensing optical system 43 are arranged in series from below. ing.

  In the present embodiment, a sample holder 71 equipped with a reactor 6 is fitted to the objective lens 8a of the microscope 8 so as to be relatively movable up and down, whereby the objective lens 8a and the reactor can be moved. The focal length with 6 samples can be adjusted. Further, when the sample holder 71 is fitted to the outer peripheral portion of the objective lens 8a, the relative vibration due to the external vibration between the objective lens 8a and the reactor 6 (specifically, the rotational vibration generated by the rotation of the rotating disk 4). The displacement is minimized. Therefore, the microscope 8 can always stably capture the sample state in the reactor 6 as a clear image without blurring, and can accurately and reliably observe the state of the sample in the separation process or the synthesis process. Become. The objective lens 8a has a field stop system 72 and an aperture stop system 73 arranged in series from below.

  Therefore, the excitation light from the illuminating device 41 is stopped by the stop system 42, condensed by the condensing optical system 43, enters the objective lens 8 a, passes through the dichroic beam splitter 14, and is back-focused by the objective lens 8 a ( Focused in the vicinity of the rear focal plane). When the sample is irradiated with the excitation light by the objective lens 8a, the reflected light (fluorescence) of the specific substance returns to the dichroic beam splitter 14, and only the light having a wavelength of 505 nm or more derived from the fluorescence emission of the specific substance. Is reflected and sent to the CCD camera 10 which is an imaging device.

  In the present embodiment, a camera image (captured image) of the microscope image captured by the CCD camera 10 is wirelessly transmitted to the rotating disk 4 in real time as well as the CCD camera 10 that captures the above-described microscope image as a movie. A video wireless transmission device 12 is attached. In addition, an excitation light removal filter 46 that removes excitation light included in the fluorescent light from the reflected light from the dichroic beam splitter 14 is disposed between the dichroic beam splitter 14 and the CCD camera 10.

  As described above, in the centrifugal microscope C according to the present embodiment, the reactor 6 is supported above the turntable 4, the illumination device 41 is disposed below the turntable 4, and the sample from the illumination device 41 to the reactor 6 is provided. The excitation light can be raised toward the target. Therefore, the illumination device 41 is disposed below the turntable 4, while the other devices are provided on the turntable 4, thereby facilitating access to the sample and the operation between the objective lens 8a and the sample. Since the distance can be shortened and the aperture ratio of the objective lens 8a can be improved, bright observation can be achieved and the apparatus can be made compact. Further, by disposing an apparatus other than the illumination device 41 while being inverted on the rotating disk 4, the lens barrel 8b can be fixed to the rotating disk 4, and vibrations of the optical system can be reduced.

  In addition, the centrifugal microscope C according to the present embodiment has an effect that the lens can be brought close to the sample and can be measured with a high aperture ratio (brightness) while facilitating access to the sample by using the inverted microscope other than the fluorescence observation. At the same time, the resolution can be improved.

  In the centrifugal microscope C of the present embodiment, a video signal is transmitted to the outside or power is supplied by providing a slip ring as in the second embodiment. In this case, contactless transmission / reception can be performed optically by combining a non-contact rotary connector (for example, model 331 manufactured by Melco Tac) using a liquid metal (such as mercury) as a conductive carrier, or a combination of a light emitting element and a light receiving element. A video signal is transmitted using a non-brush type slip ring such as a rotary connector (for example, rotary link connector Type 2 manufactured by Nippon Marco Co., Ltd.).

  By using such a slip ring, it is less affected by external radio waves and has less noise than wireless transmission. In addition, it is not necessary to install a transmitter or antenna for video transmission on the microscope, and it is not necessary to supply power to them, so that the structure of the apparatus can be simplified. Furthermore, no receiver is required, and the system is compact. This is true of the entire centrifugal microscope (common to all embodiments).

  Further, when applied to an inverted centrifugal microscope, it has been necessary to install a transmitting antenna at the center of rotation, but this is not necessary. Therefore, the Z-axis stage for moving the sample up and down can be disposed near the center of the apparatus, and the centrifugal force applied to the stage can be reduced, so that the vibration of the sample holder can be reduced. In addition, since the distance to the sample holder can be shortened, the sample stage can be smoothly moved up and down for focus adjustment. In the previous type, the inner surface of the sample stage may interfere with the lens and obstruct smooth vertical movement. This time, the guide bush is further attached so that the alignment between the sample holder and the objective lens due to elastic deformation of the member is difficult to occur.

[Fourth Embodiment]
FIG. 12 is a schematic diagram showing an optical path of a centrifugal microscope according to the fourth embodiment of the present invention. The overall configuration of the centrifugal microscope of the present embodiment is substantially the same as that of the above-described third embodiment, and will be described with reference to FIG. 10 and members having the same functions as those described in this embodiment. Are denoted by the same reference numerals, and redundant description is omitted.

  In the fourth embodiment, as shown in FIG. 13, the centrifugal microscope D is an inverted centrifugal microscope, and the main configuration is the same as that of the third embodiment. In this Embodiment, the microscope 8 is being fixed to the predetermined position (outer peripheral part) of the turntable 4 so that the state of the sample in the reactor 6 can be observed. An imaging device (CCD camera) 10 for taking a sample microscopic image in the reactor 6 captured by the microscope 8 is attached to the turntable 4. In this case, the optical path of the microscope 8 is partially refracted by the dichroic beam splitter 14 with respect to the surface 4a of the rotating disk 4 at a predetermined angle, and the CCD camera 10 receives the light of the microscope 8 refracted at the predetermined angle. It is positioned in the vicinity of the rotation center of the turntable 4 so that the above-described microscope image can be taken on the road. An excitation light removal filter 46 is disposed between the dichroic beam splitter 14 and the CCD camera 10.

  In addition, a second imaging device (CCD camera) 82 for taking a microscopic image of the sample state in the reactor 6 captured by the microscope 8 is attached to the turntable 4. In this case, a second dichroic beam splitter 81 is provided between the dichroic beam splitter 14 and the CCD camera 10. The optical path of the microscope 8 is refracted twice by the dichroic beam splitters 14 and 81, and the CCD camera 82 can take the above-described microscope image on the optical path of the microscope 8 refracted twice. It is positioned in the vicinity of the rotation center of the turntable 4. An excitation light removal filter 83 is disposed between the dichroic beam splitter 82 and the CCD camera 81.

  Therefore, the excitation light m from the illumination device 41 is stopped by the stop system 42, is collected by the condensing optical system 43, enters the objective lens 8a, passes through the dichroic beam splitter 14, and is back-focused by the objective lens 8a. Focused near (rear focal plane). When the sample is irradiated with the excitation light by the objective lens 8a, the reflected light (fluorescence) of the specific substance returns to the dichroic beam splitter 14, and light having a wavelength of 505 nm or more derived from the fluorescence emission n of the specific substance. Only reflect. Then, the fluorescence emission n of this specific substance is split into two predetermined wavelengths by the dichroic beam splitter 81, the light of one wavelength is sent to the CCD camera 10, and the light of the other wavelength is sent to the CCD camera 82. Sent.

  Thus, in the centrifugal microscope D of the present embodiment, when performing fluorescence observation, it is possible to simultaneously observe not only the fluorescence image in the 1 wavelength band but also the fluorescence image in the 2 wavelength band. In this case, a second dichroic beam splitter 81 is provided between the dichroic beam splitter 14 and the CCD camera 10, and light refracted by the dichroic beam splitter 81 is taken into the CCD camera 82. As a result, the light split by the first dichroic mirror 14 can be further split by the second dichroic mirror 81 to enable simultaneous observation of two wavelength bands. For example, when measuring cells, it is possible to observe a specific site in green by GFP and to observe an autofluorescence image of intracellular substance in red. In this case, the light split by the first dichroic mirror 14 may be split into a plurality of light beams to enable simultaneous observation over three wavelength bands.

[Fifth Embodiment]
FIG. 14 is a longitudinal sectional view of an essential part of a centrifugal microscope according to a fifth embodiment of the present invention, and FIGS. 15A and 15B are diagrams illustrating an excitation light removing filter in the centrifugal microscope according to the fifth embodiment. It is a graph showing an absorption wavelength area. In addition, the same code | symbol is attached | subjected to the member which has the same function as what was demonstrated in embodiment mentioned above, and the overlapping description is abbreviate | omitted.

  The configuration of the fifth embodiment is substantially the same as that of the centrifugal microscope A of the first embodiment. The condensing optical system is different and an excitation light removal filter 94 is added. In the fifth embodiment, as shown in FIG. 14, in the centrifugal microscope A, the microscope 8 is provided with a predetermined illumination device 41 above the lens barrel 8b, and illuminates the sample from above. So that it can be observed. In addition, the lens barrel 8b is positioned between the illumination device 41 and the mirror 14, and a diaphragm system 42 and a condensing optical system 91 are arranged in series from above.

  The condensing optical system 91 condenses the illumination light of the illumination device 41 that has passed through the aperture system 42 and transmits it to the mirror 14. In this embodiment, the condensing optical system 91 includes two condensing lenses 92 and 93 arranged in series in the irradiation direction of the irradiation light from the light source 41.

  In the condensing optical system 91, the condensing lens 92 disposed above is a plano-convex lens, for example, using a TECH SPEC φ12 plano-convex lens (PCX) (focal length 12 to 100 mm) manufactured by Edmund Optics. Yes. The condensing lens 93 disposed below is a biconvex lens, and for example, a TECH SPEC φ12 biconvex lens (DCX) (focal length 12 to 100 mm) manufactured by Edmund Optics Corporation is used.

  In this case, the two condensing lenses 92 and 93 have different refractive indexes, and the irradiation distance from the light source 41 is refracted in two stages so that the focal length can be shortened while maintaining the observation accuracy. The condenser lens 92 is a plano-convex lens and the condenser lens 93 is a biconvex lens, so that irradiation light can be received in as large an area as possible and can be uniformly condensed at an appropriate position.

  Here, the condensing optical system 91 is provided between the aperture system 42 and the dichroic beam splitter 14, but the present invention is not limited to this position. For example, the condenser lens 92 may be disposed between the aperture system 42 and the dichroic beam splitter 14, and the condenser lens 93 may be disposed between the dichroic beam splitter 14 and the objective lens. The number of condensing lenses constituting the condensing optical system 91 is not limited to two, and may be three or more depending on the type of the selected lens.

  Further, excitation that removes excitation light having a predetermined wavelength from excitation light emitted from the light source 41 between the light source 41 and the reactor, specifically, between the light source 41 and the diaphragm system 42. A light removal filter 94 is provided. The excitation light removal filter 94 removes an excitation light component in a wavelength range intended for the fluorescence wavelength region included in the excitation light from the light source 41. Although not shown, an excitation light removal filter 46 for removing excitation light contained in the fluorescence is arranged between the dichroic beam splitter 14 and the CCD camera 10 as in the first embodiment (see FIG. 4). It is installed.

In the present embodiment, two excitation light removal filters 94 and 46 are disposed between the light source 41 and the CCD camera 10. Here, the role of the excitation light removal filters 94 and 46 will be described. When only one excitation light removal filter 46 is used, in the case shown in FIG. 15A, when the predetermined wavelength region α is absorbed by the excitation light removal filter 46, the excitation light λ _{1 in the} wavelength region represented by oblique lines remains. In this case, when observing a substance having a high fluorescence emission intensity such as a fluorescent bead in the solution, the observation accuracy is not greatly affected, but the fluorescence generation intensity of the observation target is weaker and more minute, for example, GFP In the case of observing an intracellular substance in which expression is expressed, there is a risk of adversely affecting observation accuracy such as a decrease in contrast and insufficient resolution.

On the other hand, when two excitation light removal filters 94 and 46 are used, as shown in FIG. 15B, when the intensity of the fluorescence λ ₂ is very weak, first, the wavelength region of the fluorescence λ ₂ is obtained by the excitation light removal filter 94. The excitation light λ ₁ in the wavelength range including is absorbed, and preferably, the peak portion β of the intensity of the excitation light λ ₁ is transmitted and the other wavelength range is absorbed. Next, when the predetermined wavelength region α is absorbed by the excitation light removal filter 46, the excitation light λ ₁ is almost absorbed. When the intensity of the excitation light λ ₁ of the absorbed light is larger than the intensity of the fluorescence λ ₂ of the transmitted light, the amount of the remaining excitation light λ ₁ increases with the excitation light removal filter 46 alone. Even so, the light of the excitation light λ ₁ can be reliably absorbed, and a substance such as GFP or protein whose observation object is smaller than cells or glass beads can be observed with high accuracy. By combining these two excitation light removal filters 94 and 46, even when the intensity of the fluorescent light λ ₂ is very weak, it can be transmitted appropriately, and it is a large size used in a general microscope. No light source (for example, a high-pressure mercury lamp, a xenon lamp, a highly integrated LED, or a large laser light source) can be used, and fluorescence observation can be performed with a small light source. Therefore, the microscope barrel 8b can be simplified, the centrifugal rotation load can be further reduced, and a stable observation image can be obtained.

  In this case, the excitation light removal filters 94 and 46 can remove excitation light in different wavelength regions. Specifically, for example, the excitation light removal filter 46 is an absorption filter that transmits only the light of a predetermined value or more out of the reflected light. For example, by using Semlock FF01-512 / 630-25, Only light of about 25 nm width) is transmitted. Further, the removal filter 94 is an absorption filter that transmits only light in a predetermined wavelength region of the excitation light. For example, using the FF01-470 / 22-25 manufactured by Semlock, the removal filter 94 transmits only light of 450 nm to 495 nm. .

  Here, an excitation light removal filter 94 on the short wavelength side is provided between the light source 41 and the dichroic beam splitter 14, and an excitation light removal filter 46 on the long wavelength side is provided between the dichroic beam splitter 14 and the CCD camera 10. These are not limited to this position. However, in order to avoid interference with the excitation light irradiated to the sample (reactor 6), the excitation light removal filter 46 on the long wavelength side refracts the observation light by the dichroic beam splitter 14, and the optical path of the irradiated excitation light It is desirable to provide after separation. For the same reason, it is desirable to provide the excitation light removal filter 94 on the short wavelength side between the light source 41 and the dichroic beam splitter 14.

  In the centrifugal microscope A of the present embodiment, the excitation light from the illumination device 41 is narrowed by passing through the diaphragm system 42, and the diaphragm light is condensed by the condensing optical system 91 and enters the objective lens. . In this case, the aperture light is adjusted by the condensing optical system 91 so that it passes through the dichroic beam splitter 14 and is condensed near the back focus (rear focal plane) of the objective lens. Then, when the sample is irradiated with the excitation light by the objective lens, the reflected light (fluorescence) generated from the specific substance returns to the dichroic beam splitter 14, and only the light derived from the fluorescence emission of the specific substance is refracted. And sent to a CCD camera as an imaging device.

  As described above, in the centrifugal microscope A according to the present embodiment, the condensing optical system 91 is disposed between the light source 41 and the dichroic beam splitter 14, and the condensing optical system 91 is connected in series in the irradiation direction of the excitation light. Two condensing lenses 92 and 93 are provided. In general, in order to reduce the focal length of a condenser lens, it is necessary to increase the refractive index of the lens or to reduce the radius of curvature. For this purpose, it is conceivable to use a lens made of a special material, but this leads to an increase in cost. Further, although a lens having a small curvature radius such as a ball lens may be employed, various monochromatic aberrations of the lens become large, and there is a possibility that the observation accuracy such as a reduction in resolution and distortion of the image may be caused. These aberrations are caused by interference between the light transmitted through the center of the lens and the light transmitted through the end of the lens. In a general microscope, an iris is provided to limit the light transmission area to the vicinity of the center of the lens. Is done. However, in this case, the light source needs to be strengthened (high output) to ensure the brightness of the image, which is inappropriate for the centrifugal microscope in which the power source capacity, installation space, and the like are limited. Therefore, in this embodiment, by using the plurality of condensing lenses 92 and 93, the distance from the light source 41 to the focal length can be shortened, and the apparatus can be made compact. As a result, the apparatus can be reduced in weight, the centrifugal force acting on the microscope at the time of rotation can be reduced, and vibration can be reduced to improve the observation accuracy. Furthermore, it is not necessary to use a lens made of a special material, and the production cost can be reduced.

  In addition, an excitation light removal filter 94 that removes light in a predetermined wavelength region from the excitation light is disposed between the light source 41 and the condensing optical system 91, and reflected between the dichroic beam splitter 14 and the CCD camera 10. An excitation light removal filter 46 that removes light in a predetermined wavelength region from the light is disposed, and each excitation light removal filter 94, 46 removes light in a different wavelength region. Therefore, the excitation light in the wavelength range that enters the wavelength region of the fluorescence is removed from the excitation light emitted from the light source and the fluorescence reflected by the dichroic beam splitter 14, and a clearer image can be obtained. A highly accurate fluorescent image can be obtained regardless of the form (wavelength characteristics, intensity) and the observation object.

[Sixth Embodiment]
FIG. 16 is a graph showing the numerical aperture and working distance of the condensing optical system in the centrifugal microscope according to the sixth embodiment of the present invention. The overall configuration of the centrifugal microscope according to the present embodiment is substantially the same as that of the first embodiment described above, and will be described with reference to FIG. 4 and a member having the same function as that described in the present embodiment. Are denoted by the same reference numerals and redundant description is omitted.

  In the sixth embodiment, a centrifugal microscope is employed in which an epi-illumination device that illuminates the sample of the reactor from above is added. In this epi-illumination type centrifugal microscope, a working space is required between the objective lens and the reactor, but this space is limited due to the structure. For this reason, when selecting an objective lens, it is desirable to use a lens having a large aperture ratio (bright) while maintaining a certain working distance. In the present embodiment, as shown in FIG. 4, the microscope 8 of the centrifugal microscope A is fixed at a predetermined position on the rotating disk 4 and includes an objective lens 8 a that captures the state of the sample in the reactor 6, and an objective lens 8 a. It has a lens barrel 8b in which an optical path for transmitting the captured microscopic image to the imaging device 10 is formed. Further, the microscope 8 is provided with an illumination device 41, a diaphragm system 42, a condensing optical system 43, and a dichroic beam splitter 14 in a lens barrel 8b as fluorescence observation means capable of fluorescence observation of a sample of the reactor 6. Configured.

  In the microscope 8 thus configured, the objective lens 8a has a numerical aperture in the range of 0.3 to 1.3 and a working distance in the range of 0.2 to 18.0 mm. That is, the function as the objective lens 8a is satisfactorily exhibited by optimizing the relationship between the numerical aperture and the working distance. For example, as shown in FIG. 16, in the case of the region c in which the numerical aperture is increased and the working distance is shortened, the resolution is improved, but the space near the reactor 6 is reduced. It will decline. In addition, since there are covers and protrusions on the sample such as a preparation and it is difficult to bring the lens close to the sample, there arises a problem that the sample may not be observed. On the other hand, in the region a where the numerical aperture is lowered and the working distance is lengthened, the space near the reactor 6 becomes sufficient and workability such as sample exchange is improved, but the resolution is lowered.

  Therefore, in the present embodiment, the objective lens 8a is set to have a numerical aperture in the range of 0.3 to 1.3 and a working distance in the range of 0.2 to 18.0 mm. In this case, the objective lens 8a is preferably set to a numerical aperture of 0.5 to 0.55 and a working distance of 8.3 to 10.6 mm.

  Thus, in the epi-illumination type centrifugal microscope A according to the present embodiment, the numerical aperture of the objective lens 8a is in the range of 0.3 to 1.3, and the working distance is in the range of 0.2 to 18.0 mm. Preferably, the objective lens 8a is set to have a numerical aperture of 0.5 to 0.55 and a working distance of 8.3 to 10.6 mm. Therefore, by optimizing the relationship between the numerical aperture and the working distance, it is possible to achieve both high resolution of the fluorescent image and high operability of the apparatus.

[Seventh Embodiment]
FIG. 17 is a longitudinal sectional view of an essential part of a centrifugal microscope according to the seventh embodiment of the present invention, and FIGS. 18A and 18B are schematic views showing modifications of the centrifugal microscope according to the seventh embodiment. FIG. In addition, the same code | symbol is attached | subjected to the member which has the same function as what was demonstrated in embodiment mentioned above, and the overlapping description is abbreviate | omitted.

  In the seventh embodiment, as shown in FIG. 17, the microscope 8 of the centrifugal microscope has an objective lens 8 a that captures the state of the sample in the reactor 6. Further, the microscope 8 includes an illumination device, a diaphragm system, a condensing optical system, and a dichroic beam splitter as fluorescence observation means capable of fluorescence observation of the sample in the reactor 6.

  In the microscope 8 configured in this manner, the fluorescence observation fluid is filled between the objective lens 8a and the reactor 6, and a cap 101 for preventing leakage and scattering of the fluorescence observation fluid is disposed. Has been. The cap 101 has a cylindrical shape, and a lower end portion is placed on the upper surface of the reactor 6. Further, the cap 101 is formed with a ring-shaped tapered surface 101a having a lower height on the inner peripheral side at the upper end portion, and this tapered surface 101a is parallel to the inclined surface forming the truncated cone shape of the objective lens 8a. They are arranged with a predetermined gap or in close contact with each other. The space surrounded by the objective lens 8a, the cap 101, and the reactor 6 is filled with a fluorescence observation fluid.

  Note that this fluorescence observation fluid has a higher refractive index than air, for example, oil immersion oil, which improves resolution and contrast. The cap 101 is preferably made of an elastic material such as silicon rubber (PDMS: polydimethylsiloxane), and is appropriately interposed between the objective lens 8a and the reactor 6 and filled inside by a centrifugal force. Suppresses leakage and scattering. In this case, it is preferable to color so that excitation light is not scattered or external disturbance light is not transmitted.

  The cap 101 is not limited to this shape. In the modified example of the centrifugal microscope of the seventh embodiment, as shown in FIG. 18A, the microscope 8 is filled with the fluorescence observation fluid between the objective lens 8a and the reactor 6, as shown in FIG. A cap 102 is provided to prevent leakage and scattering of the fluorescence observation fluid. The cap 102 has a cylindrical shape that is bent so that the upper and lower intermediate portions are thin, and the lower end portion is placed on the upper surface of the reactor 6. Further, the cap 102 is formed with a ring-shaped tapered surface 102a having a lower height on the inner peripheral side at the upper end portion, and this tapered surface 102a is parallel to the inclined surface forming the truncated cone shape of the objective lens 8a. They are arranged with a predetermined gap or in close contact with each other. The space surrounded by the objective lens 8a, the cap 102, and the reactor 6 is filled with a fluorescence observation fluid.

  Further, as shown in FIG. 18-2, the microscope 8 is filled with the fluorescence observation fluid between the objective lens 8a and the reactor 6, and prevents leakage and scattering of the fluorescence observation fluid. A cap 103 is provided. The cap 103 has a cylindrical shape that is curved so that the upper and lower intermediate portions are thin, and the lower end portion is placed on the upper surface of the reactor 6. Further, the cap 103 is disposed so that the inner peripheral surface 103a at the upper end portion is in close contact with the inclined surface forming the truncated cone shape of the objective lens 8a. The space surrounded by the objective lens 8a, the cap 103, and the reactor 6 is filled with a fluorescence observation fluid.

  Furthermore, the cap is not limited to such a configuration. For example, a labyrinth seal (branched at the end) is used so as to ensure slidability and adhesion to the objective lens 8a and the reactor 6. It may be.

  As described above, in the centrifugal microscope of the present embodiment, the caps 101, 102, and 103 that fill the fluorescent observation fluid between the objective lens 8a and the reactor 6 and prevent the leakage and scattering of the fluorescent observation fluid are provided. It is arranged. Accordingly, it is possible to obtain a highly accurate fluorescent image with the fluorescence observation fluid, and it is possible to prevent leakage and scattering of the fluorescence observation fluid with the caps 101, 102, and 103, thereby improving the reliability of the apparatus. Can do.

  Note that the centrifugal microscope of the present invention can observe a micro sample while applying centrifugal hypergravity, as shown in the above-described embodiments. By utilizing this, for example, the adhesion of a micro object such as a cell to the substrate surface can be quantitatively measured. Conversely, as the characteristic evaluation of the substrate surface, the spread of liquid (wetability) under centrifugal gravity can be evaluated. In addition to gravity in one direction, it is possible to control the spatial position of fine particles using Coriolis force by combining acceleration and deceleration. Needless to say, it is effective for analyzing the behavior of various cells and bacteria under gravity. In particular, in applications intended for biomolecules such as cells, the localization and expression state of a specific protein can be observed by the fluorescence observation function, which is effective.

  As described above, the best mode for carrying out the present invention has been described. However, the present invention is not limited to these, and various modifications are possible within the scope of the technical idea of the present invention.

2 Rotating shaft 4 Rotating disk 6 Reactor 8 Microscope 8a Objective lens 10, 82 Imaging device (CCD camera)
12 Video wireless transmission device 14, 81 Mirror, dichroic beam splitter (spectrometer)
Reference Signs List 16 Fixed component 18 Antenna 20 Z-axis guide 22 Illumination device 24 Power supply device 28 Spindle unit 30 Balance weight 41 Illumination device (light source)
42 Aperture system 43, 91 Condensing optical system 46, 83, 94 Excitation light removal filter 60 Sample mounting part A, B, C, D Centrifugal microscope 101, 102, 103 Cap λ1 Waveform of excitation light λ2 Waveform of fluorescence α Excitation light Wavelength absorption range of removal filter 46 β Wavelength transmission range of excitation light removal filter 94 (peak portion of excitation intensity)

Claims (9)

A rotatable turntable,
A reactor disposed on the outer peripheral side of the rotating disk and containing a sample;
An imaging device disposed in the center of the turntable and capable of taking an image of the sample state of the reactor by rotating together with the turntable;
In a centrifugal microscope comprising:
Provide fluorescence observation means capable of fluorescence observation of the sample of the reactor,
The fluorescence observation means includes
A light source using a semiconductor laser disposed on the rotating plate;
An objective lens for irradiating the sample of the reactor with excitation light from the light source;
A spectroscope that transmits the excitation light from the light source, and refracts the fluorescence generated from the sample of the reactor with the excitation light and refracts it to the imaging device;
Have
Between the spectroscope and the imaging device, an excitation light removal filter that removes excitation light from reflected light by the spectrometer is disposed.
A centrifugal microscope characterized by that.   The reactor is supported on the turntable, the light source is disposed above the turntable, and excitation light can be incident on the sample of the reactor from the light source. The centrifugal microscope according to 1.   The reactor is supported above the turntable, the light source is disposed below the turntable, and excitation light can be emitted from the light source toward the sample of the reactor. The centrifugal microscope according to claim 1.   The light source, the spectroscope, the objective lens, and the reactor are arranged in series along a rotation axis direction of the rotating disk, and the spectroscope, the excitation light removing filter, and the imaging device are connected to the rotating disk. The centrifugal microscope according to any one of claims 1 to 3, wherein the centrifugal microscope is arranged in series along a radial direction.   The centrifugal microscope according to any one of claims 1 to 4, wherein a diaphragm system and a condensing optical system are disposed between the light source and the spectroscope.   A condensing optical system is disposed between the light source and the objective lens, and the condensing optical system includes a plurality of condensing lenses arranged in series in the irradiation direction of the excitation light. Item 6. The centrifugal microscope according to any one of Items 1 to 5.   Between the light source and the spectroscope, one or more excitation light removal filters for removing excitation light in a range that enters the fluorescence wavelength region from excitation light irradiated from the light source are disposed. The centrifugal microscope according to any one of claims 1 to 6, wherein:   8. The fluorescent observation fluid is filled between the objective lens and the reactor, and a cap for preventing leakage and scattering of the fluorescence observation fluid is disposed. The centrifugal microscope according to any one of the above.   9. The objective lens according to claim 1, wherein a numerical aperture is set in a range of 0.3 to 1.3 and a working distance is set in a range of 0.2 to 18.0 mm. The centrifugal microscope according to one.

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