JP2004309270A - Microchemical system - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a microchemical system in which both detecting methods of photothermal conversion spectroscopic analysis and LIF analysis can be utilized. <P>SOLUTION: A microchemical system 1 is provided with a microchemical system chip 20 enabling sample solution to flow in a channel 204; an irradiation section 1a being shared by the photothermal conversion spectroscopic analysis and the LIF analysis; a light receiving section 1b used for the photothermal conversion spectroscopic analysis; and a light receiving section 1c used for the LIF analysis. The light receiving section 1c includes an optical fiber 302 which receives fluorescent light in a direction normal to the optical axis of exciting light emitted from the irradiation section 1a; and a fluorescence condenser lens 301 which is coupled to a leading edge of the optical fiber 302 and embedded in the microchemical system chip 20. <P>COPYRIGHT: (C)2005,JPO&NCIPI

Description

[0001]
TECHNICAL FIELD OF THE INVENTION
The present invention relates to a microchemical system, and more particularly to a microchemical system used for photothermal conversion spectroscopy.
[0002]
[Prior art]
Conventionally, integrated technologies for performing chemical reactions in a minute space have attracted attention from the viewpoints of high-speed chemical reactions, reactions in minute amounts, on-site analysis, etc. I have.
[0003]
A microchemical system using a glass substrate, etc., as one of the chemical reaction integration technologies, is a method for mixing, reacting, separating, extracting, detecting, etc. of samples in a fine channel created on a small glass substrate, etc. It aims to be able to demonstrate the function of. Examples of reactions performed in a microchemical system include diazotization, nitration, and antigen-antibody reactions, and examples of extraction and separation include solvent extraction, electrophoretic separation, and column separation.
[0004]
Among the above functions, an electrophoresis apparatus for analyzing only a trace amount of protein, nucleic acid, or the like has been proposed as one intended only for separation (for example, see Patent Document 1). The electrophoresis apparatus includes a plate-like member with a flow path formed of two glass substrates bonded to each other. Since this member is plate-shaped, it is less likely to be damaged than a glass capillary tube having a circular or square cross section, and is easy to handle.
[0005]
In microchemical systems, since the amount of sample is very small, a highly sensitive detection method is indispensable. This highly sensitive detection method is based on the thermal lens generated by the light absorption of a submerged sample in a fine channel. The establishment of the photothermal conversion spectroscopy utilizing the effect has opened the way to practical use (for example, see Patent Document 2).
[0006]
In photothermal conversion spectroscopy, when a sample is condensed and irradiated with light, the solvent locally rises in temperature due to the heat energy released due to the light absorption of the solute in the sample, and the refractive index changes. As a result, an analysis method utilizing the photothermal conversion effect that a thermal lens is formed is used.
[0007]
FIG. 3 is an explanatory diagram of the principle of the thermal lens.
[0008]
In FIG. 3, when the excitation light is condensed and irradiated on the very small sample through the objective lens of the microscope, a photothermal conversion effect is induced. For many substances, the refractive index decreases with increasing temperature. Therefore, in the sample irradiated with the excitation light, the refractive index decreases more as the temperature rises closer to the light-collecting center, and the refractive index decreases as the temperature rises closer to the periphery due to thermal diffusion. Is small. Optically, this refractive index distribution has exactly the same effect as a concave lens, so this effect is called a thermal lens effect, and the magnitude of the effect, that is, the power of the concave lens is proportional to the light absorption of the sample. On the contrary, when the refractive index increases in proportion to the temperature, the same effect as that of the convex lens is obtained.
[0009]
As described above, the photothermal conversion spectroscopy is for observing the diffusion of heat, that is, the change in the refractive index, and is therefore suitable for detecting the concentration of an extremely small sample.
[0010]
A laser-induced fluorescence (LIF: Laser Induced Fluorescence) analysis is another highly sensitive detection method other than the photothermal conversion spectroscopy. The LIF analysis is a method in which a target fluorescent molecule is electronically excited by a laser, and the fluorescence generated when the target fluorescent molecule falls to the ground level is measured. Since the resonance transition between energy levels is used, the probability of its excitation is high, and extremely sensitive detection is possible. For this reason, LIF analysis is also suitable for detecting the concentration of an extremely small sample.
[0011]
As an example of this detection method, one that measures the fluorescence of a sample flowing in a fine channel formed on a small glass substrate or the like is disclosed (for example, see Non-Patent Documents 1 and 2). In the methods disclosed in these articles, the excitation light is guided to a sample flowing in the flow path by an optical fiber or a waveguide embedded in a small glass substrate or the like up to the vicinity of the flow path.
[0012]
In a conventional microchemical system, a plate member with a flow path is disposed below an objective lens of a microscope, and excitation light having a predetermined wavelength output from an excitation light source is incident on the microscope, and the objective lens of the microscope uses the excitation light. The sample in the analysis channel of the plate member with the channel is focused and irradiated. A thermal lens is formed around the converging irradiation position of this converging irradiation.
[0013]
On the other hand, the detection light output from the detection light source and having a wavelength different from the excitation light is incident on the microscope, and the detection light emitted from the microscope is condensed and irradiated on the thermal lens formed on the sample by the excitation light, and the sample is irradiated. Transmit and diverge or condense. The light diverged or condensed from the sample and emitted is signal light, and the signal light is detected by the detector via only the condenser lens and the filter or only the filter. The intensity of the signal light detected by this detector depends on the thermal lens formed on the sample.
[0014]
As described above, in the photothermal conversion spectrometer described above, the thermal lens is formed at the position where the excitation light is condensed and irradiated (hereinafter, referred to as “focal position”), and the change in the refractive index of the thermal lens formed is as follows. The wavelength is detected by detection light different from the excitation light.
[0015]
[Patent Document 1]
JP-A-8-178897
[Patent Document 2]
JP-A-10-232210
[Non-patent document 1]
Rev .. Sci. Instrum. , Vol. 72, No. 1, P229, 2001
[Non-patent document 2]
Anal. Chem. , Vol. 68, no. 6, P1040, 1996
[0016]
[Problems to be solved by the invention]
However, the photothermal conversion spectrometer described above is large and lacks portability due to the complicated configuration of the light source, the optical system of the measurement unit and the detection unit (photoelectric conversion unit). For this reason, when performing analysis or the like using this photothermal conversion spectroscopic analyzer, there is a problem that the operation of the place or the device is limited, and furthermore, there is a problem that the work efficiency of the user is poor.
[0017]
In addition, since the photothermal conversion spectrometer guides the excitation light and the detection light to the sample as spatial light, each component of the optical system such as a light source, a mirror, and a lens must be kept from moving during the measurement. In order to fix them, a solid surface plate is required. Further, when the optical axes of the excitation light and the detection light are shifted due to a change in environment such as temperature, a jig for adjusting the shift is required. These factors also cause the photothermal conversion spectrometer to be large and lack portability.
[0018]
In the photothermal conversion spectrometer, it is necessary that the focal position of the excitation light and the focal position of the detection light are different for the following reasons.
[0019]
FIGS. 4A and 4B are explanatory diagrams of the formation position of the thermal lens and the focus position of the detection light in the optical axis direction (Z direction) of the excitation light. FIG. 4A shows a case where the objective lens has chromatic aberration, and FIG. Indicates a case where the objective lens has no chromatic aberration.
[0020]
When the objective lens 130 has chromatic aberration, as shown in FIG. 4A, the thermal lens 131 is formed at the focus position 132 of the excitation light, and the focus position 133 of the detection light is the focus of the excitation light by ΔL. Since the position 132 deviates from the position 132, a change in the refractive index of the thermal lens 131 can be detected as a change in the focal length of the detection light by the detection light. On the other hand, when the objective lens 130 does not have chromatic aberration, as shown in FIG. 4B, the focal position 133 of the detection light is substantially equal to the position of the thermal lens 131 formed at the focal position 132 of the excitation light. As a result, the detection light is not deflected by the thermal lens 131, and the change in the refractive index of the thermal lens 131 cannot be detected.
[0021]
However, the objective lens of the microscope is usually manufactured so as not to have chromatic aberration. Therefore, for the above-described reason, the focus position 133 of the detection light is different from that of the thermal lens 131 formed at the focus position 132 of the excitation light. The position almost coincides with the position (FIG. 4B), and the change in the refractive index of the thermal lens 131 cannot be detected. For this reason, each time the measurement is performed, the position where the thermal lens is formed is shifted from the focus position 133 of the detection light as shown in FIGS. 5A and 5B, or as shown in FIG. It is necessary to shift the focal position 133 of the detection light from the thermal lens 131 by making the detection light incident at a slight angle using a lens (not shown), and this also has a problem that the work efficiency of the user is poor. is there.
[0022]
Since LIF analysis is also suitable for detecting the concentration of a very small sample, it may be more useful to use LIF analysis as a means of detecting a microchemical system depending on the sample to be measured. Since there is no microchemical system capable of performing both photothermal conversion spectroscopy, there is a problem in that separate systems must be provided for each. In addition, a sample may contain a fluorescent substance that can be detected by LIF analysis and a non-fluorescent substance that cannot be detected. In such a case, usually, the fluorescent substance is first detected by LIF analysis, and then the fluorescent substance is detected. A very laborious operation of detecting a non-fluorescent substance by photothermal conversion spectroscopy using a different apparatus was required.
[0023]
An object of the present invention is to provide a microchemical system in which both detection methods of photothermal conversion spectroscopy and LIF analysis can be used.
[0024]
[Means for Solving the Problems]
In order to achieve the above object, the microchemical system according to claim 1 irradiates a sample with excitation light and detection light for photothermal conversion spectroscopy via an irradiation lens, and irradiates the sample with the excitation light. A detection means for detecting the detection light transmitted through the thermal lens generated in the sample; and a detection means for detecting fluorescence emitted from the sample by irradiation of the excitation light. It is characterized by.
[0025]
According to the microchemical system according to claim 1, in the microchemical system including an irradiation unit and a detection unit for photothermal conversion spectroscopy, another detection unit that detects fluorescence emitted from a sample irradiated with the excitation light is provided. , It is possible to provide a microchemical system in which both detection methods of photothermal conversion spectroscopy and laser-induced fluorescence analysis can be used. As a result, for a sample in which a fluorescent substance and a non-fluorescent substance are mixed, the fluorescent substance and the non-fluorescent substance can be detected at the same time, the work efficiency of the user can be improved, and the reaction process and the yield can be identified. For a sample consisting only of a fluorescent substance, photothermal conversion spectroscopy and laser-induced fluorescence analysis can be performed simultaneously, and by correlating the results of the two, separation from noise is facilitated to improve measurement sensitivity. be able to.
[0026]
A microchemical system according to a second aspect is characterized in that, in the microchemical system according to the first aspect, the other detecting means is arranged at a position not on the optical axis.
[0027]
According to the microchemical system according to the second aspect, since the other detection means is arranged at a position not on the optical axis of the excitation light, even if the photothermal conversion spectroscopy analysis and the laser-induced fluorescence analysis are performed simultaneously, It is possible to prevent excitation light detected as noise from being incident on the detection means, and to prevent a decrease in measurement sensitivity of laser-induced fluorescence analysis. Preferably, the other detecting means is arranged at a position orthogonal to the optical axis of the excitation light.
[0028]
The microchemical system according to claim 3, wherein the microchemical system according to claim 2, further comprising a transparent substrate having a flow path for accommodating the sample, wherein the other detection unit includes a fluorescence focusing lens, The fluorescence condensing lens is buried in a peripheral portion of the flow path in the transparent substrate or disposed on an outer surface of the substrate.
[0029]
According to the microchemical system according to claim 3, the transparent chemical substrate includes a transparent substrate having a channel for accommodating a sample, and the other detecting means includes a fluorescent light focusing lens, and the fluorescent light focusing lens is provided in the transparent substrate. Embedded in the peripheral portion of the flow path or disposed on the outer surface of the substrate, more fluorescence emitted from the sample flowing through the flow path can be received, and the measurement sensitivity of the laser-induced fluorescence analysis can be improved.
[0030]
A microchemical system according to a fourth aspect is the microchemical system according to any one of the first to third aspects, further comprising one path for propagating the excitation light and the detection light to the irradiation lens.
[0031]
According to the microchemical system of the fourth aspect, since one path for transmitting the excitation light and the detection light to the irradiation lens is provided, the excitation light and the detection light are always coaxial, and the optical axis alignment is not required. it can.
[0032]
A microchemical system according to a fifth aspect of the present invention is the microchemical system according to the fourth aspect, wherein the one path comprises an optical fiber.
[0033]
According to the microchemical system according to the fifth aspect, since one path for propagating the excitation light and the detection light to the irradiation lens is formed of an optical fiber, the excitation light and the detection light can be respectively transmitted without being affected by external changes. It can propagate to the illumination lens.
[0034]
According to a sixth aspect of the present invention, in the microchemical system according to the fifth aspect, the optical fiber is a single mode fiber.
[0035]
According to the microchemical system according to claim 6, since the optical fiber constituting one path is a single mode fiber, the thermal lens generated by the excitation light becomes a small thermal lens with small aberration, Measurement sensitivity can be improved.
[0036]
The microchemical system according to claim 7, wherein the detection light has a wavelength different from the wavelength of the excitation light, and the irradiation lens has chromatic aberration. Characterized by comprising a lens having:
[0037]
According to the microchemical system according to claim 7, the detection light has a wavelength different from the wavelength of the excitation light, and the irradiation lens is formed of a lens having chromatic aberration. It can be shifted without using an optical system, and thus the microchemical system can be downsized.
[0038]
The microchemical system according to claim 8 is the microchemical system according to claim 7, wherein the irradiation lens is a rod lens.
[0039]
According to the microchemical system of the present invention, since the irradiation lens is a rod lens, it is easy to match the optical axes of the excitation light and the detection light with the optical axis of the rod lens, and the irradiation lens can be downsized. Therefore, the size of the microchemical system can be further reduced.
[0040]
According to a ninth aspect of the present invention, in the microchemical system according to the eighth aspect, the irradiation lens is a gradient index rod lens.
[0041]
According to the microchemical system according to the ninth aspect, since the irradiation lens is a gradient index rod lens, the size of the irradiation lens can be further reduced, so that the size of the microchemical system can be further reduced.
[0042]
A microchemical system according to a tenth aspect is the microchemical system according to any one of the first to ninth aspects, further comprising an excitation light output adjusting device that modulates an output of the excitation light.
[0043]
According to the microchemical system of the present invention, since the output modulator for modulating the output of the excitation light is provided, in the photothermal conversion spectroscopy, the thermal lens formed by the irradiation of the excitation light is excited so as not to be saturated. It is not necessary to install a device such as a chopper for periodically turning on and off the light irradiation, and the transmission path of the excitation light can be entirely within the optical fiber, miniaturizing the microchemical system. In addition, the excitation light can be propagated to the irradiation lens without being affected by external changes, so that the measurement sensitivity of the microchemical system can be improved.
[0044]
In order to achieve the above object, the microchemical system according to claim 11 irradiates the sample with an excitation light and a detection light for photothermal conversion spectroscopy via an irradiation lens, and irradiates the sample with the excitation light. A detection means for detecting the detection light transmitted through the thermal lens generated in the sample, wherein the detection means also detects fluorescence emitted from the sample by irradiation of the excitation light. And
[0045]
According to the microchemical system according to claim 11, in the microchemical system including the irradiation unit and the detection unit for photothermal conversion spectroscopy, the detection unit also detects fluorescence emitted from the sample by irradiation with the excitation light, The whole system can be made compact.
[0046]
The microchemical system according to claim 12 is the microchemical system according to claim 11, wherein the microchemical system further includes a detection light transmission unit that transmits only the detection light and a fluorescence transmission unit that transmits only the fluorescence. The detection light transmission means and the fluorescence transmission means are provided so as to be able to enter and exit between the detection means and the sample on the optical axis of the excitation light.
[0047]
According to the microchemical system according to the twelfth aspect, the detection light transmitting unit that transmits only the detection light and the fluorescence transmission unit that transmits only the fluorescence can be moved in and out between the detection unit and the sample on the optical axis of the excitation light. Since it is installed, when performing photothermal conversion spectroscopy, the detection light transmission means is inserted on the optical axis of the excitation light, and when performing LIF analysis, the fluorescence transmission means is inserted on the optical axis of the excitation light. The analysis contents can be switched.
[0048]
A microchemical system according to a thirteenth aspect is the microchemical system according to the twelfth aspect, wherein the microchemical system includes one path for propagating the excitation light and the detection light to the irradiation lens.
[0049]
According to the microchemical system according to the thirteenth aspect, since one path for transmitting the excitation light and the detection light to the irradiation lens is provided, the excitation light and the detection light are always coaxial, and the optical axis alignment is not required. it can.
[0050]
According to a fourteenth aspect of the present invention, in the microchemical system according to the thirteenth aspect, the one path comprises an optical fiber.
[0051]
According to the microchemical system according to claim 14, one path for transmitting the excitation light and the detection light to the irradiation lens is formed of an optical fiber, so that the excitation light and the detection light are not affected by external changes. It can propagate to the illumination lens.
[0052]
According to a fifteenth aspect of the present invention, in the microchemical system according to the fourteenth aspect, the optical fiber is a single mode fiber.
[0053]
According to the microchemical system of claim 15, since the optical fiber constituting one path is a single mode fiber, the thermal lens generated by the excitation light becomes a small thermal lens with small aberration, Measurement sensitivity can be improved.
[0054]
The microchemical system according to claim 16, wherein the detection light has a wavelength different from the wavelength of the excitation light, and the irradiation lens has chromatic aberration. Characterized by comprising a lens having:
[0055]
According to the microchemical system according to claim 16, the detection light has a wavelength different from the wavelength of the excitation light, and the irradiation lens is formed of a lens having chromatic aberration. It can be shifted without using an optical system, and thus the microchemical system can be downsized.
[0056]
A microchemical system according to a seventeenth aspect is the microchemical system according to the sixteenth aspect, wherein the irradiation lens is a rod lens.
[0057]
According to the microchemical system of the present invention, since the irradiation lens is a rod lens, it is easy to align the optical axis of the excitation light and the detection light with the optical axis of the rod lens, and the irradiation lens can be downsized. Thus, the size of the microchemical system can be further reduced.
[0058]
The microchemical system according to claim 18 is the microchemical system according to claim 17, wherein the irradiation lens is a gradient index rod lens.
[0059]
According to the microchemical system of the present invention, since the irradiation lens is a gradient index rod lens, the size of the irradiation lens can be further reduced, so that the size of the microchemical system can be further reduced.
[0060]
A microchemical system according to a nineteenth aspect is the microchemical system according to any one of the eleventh to eighteenth aspects, further comprising an excitation light output adjusting device that modulates an output of the excitation light.
[0061]
According to the microchemical system according to the nineteenth aspect, since the output chemical modulator for modulating the output of the excitation light is provided, in the photothermal conversion spectroscopy, the thermal lens formed by the irradiation of the excitation light is excited so as not to be saturated. It is not necessary to install a device such as a chopper for periodically turning on and off the light irradiation, and the transmission path of the excitation light can be entirely within the optical fiber, miniaturizing the microchemical system. In addition, the excitation light can be propagated to the irradiation lens without being affected by external changes, so that the measurement sensitivity of the microchemical system can be improved.
[0062]
BEST MODE FOR CARRYING OUT THE INVENTION
Hereinafter, a configuration of a microchemical system according to an embodiment of the present invention will be described with reference to the drawings.
[0063]
FIG. 1 is a schematic diagram showing a schematic configuration of the microchemical system according to the first embodiment of the present invention.
[0064]
In FIG. 1, a microchemical system 1 applies a photothermal conversion spectroscopic analysis to a sample solution in a flow path 204 used for mixing, searching, synthesizing, separating, extracting, detecting, etc. of a sample solution inside a microchemical system chip 20. Irradiating section 1a for irradiating the excitation light and the detection light, excitation light for laser-induced fluorescence (LIF) analysis, and receiving section for receiving the detection light transmitted through the microchemical system chip 20 in the photothermal conversion spectroscopy. 1b, and a light receiving section 1c for receiving the fluorescence emitted from the sample in the channel 204 in the LIF analysis.
[0065]
The microchemical system chip 20 is composed of glass substrates 201, 202, and 203 which are bonded in three layers. In the glass substrate 202, the above-mentioned flow path 204 through which a sample flows when mixing, searching, synthesizing, separating, extracting, detecting, or the like is formed.
[0066]
The material of the microchemical system chip 20 is desirably glass from the viewpoints of durability and chemical resistance. Further, in consideration of use for biological samples such as cells, for example, for DNA analysis, glass having high acid resistance and alkali resistance is preferable. Specifically, preferred are silicate shelf glass, soda lime glass, alumino shelf silicate glass, and quartz glass. However, an organic substance such as a plastic can be used by limiting the use.
[0067]
Examples of the adhesive for bonding the glass substrates 201, 202, and 203 include an ultraviolet-curing type, a thermosetting type, a two-part curing type acrylic or epoxy organic adhesive, and a sheetless adhesive. Further, the glass substrates 201, 202, and 203 may be fused together by heat fusion.
[0068]
The irradiation unit 1a includes an excitation light source 106 that outputs excitation light (for example, a wavelength of 658 nm) used for photothermal conversion spectroscopy and LIF analysis, and detection light (for example, wavelengths) for detecting a sample in photothermal conversion spectroscopy. 785 nm), a modulator for modulating the excitation light, a modulator for modulating the excitation light, and a light source for the excitation light 106 and a light source for the detection light 107 connected via an optical fiber, respectively. A two-wavelength multiplexing element 109 for multiplexing the excitation light from 106 and the detection light from the detection light source 107; an optical fiber 103 for propagating the multiplexed excitation light and detection light in a single mode; A lens 101 attached to the tip of 103 for irradiating the propagated excitation light and detection light to a channel 204 in the microchemical system chip 20; The consists adjusted and held to the jig 102. As facing the flow channel 204 of the microchemical system chip 20. Here, the lens 101 according to the present embodiment is composed of a gradient index rod lens.
[0069]
As described above, since both the excitation light and the detection light propagate to the lens 101 via the same path (optical fiber 103), the excitation light and the detection light are always coaxial in the photothermal conversion spectroscopy, so that the excitation light and the detection light The optical axis alignment of light can be made unnecessary, and in addition, in photothermal conversion spectroscopy analysis and LIF analysis, excitation light and detection light can propagate to the lens 101 without being affected by external changes.
[0070]
Furthermore, since the optical fiber 103 and the lens 101 are used to propagate and collect the excitation light and detection light for photothermal conversion spectroscopy and the excitation light for LIF analysis, the system is greatly simplified. As a result, the size of the microchemical system 1 can be improved, and the measurement sensitivity of the microchemical system 1 can be improved. In addition, since the excitation light for photothermal conversion spectroscopy and the excitation light for LIF analysis are both emitted from the excitation light source 106, the light source is switched and adjusted for each measurement of photothermal conversion spectroscopy and LIF analysis. There is no need to improve the work efficiency of the user. In addition, since a jig or the like necessary for switching the light source for each measurement is not required, the microchemical system 1 can be reduced in size, the optical system can be easily adjusted, and the measurement sensitivity can be improved.
[0071]
Further, since the excitation light is modulated by the modulator 108, a chopper or the like for periodically turning on and off the irradiation of the excitation light in photothermal conversion spectroscopy so that the thermal lens formed by the irradiation of the excitation light is not saturated. It is not necessary to install the device outside, and the transmission path of the excitation light can be entirely within the optical fiber 103, and the microchemical system 1 can be reduced in size, without being affected by external changes. Since the excitation light can propagate to the lens 101, the measurement sensitivity of the microchemical system 1 can be improved.
[0072]
In the LIF analysis, the measurement can be performed even if the excitation light is modulated. However, when the measurement is performed without the modulation, more fluorescence is emitted from the sample, so that the measurement sensitivity is improved. Therefore, when performing only the LIF measurement, a switch for turning off the modulator 108 may be provided.
[0073]
The lens 101 is connected to an optical fiber 103 that transmits the light in a single mode on the incident side (upper side in the drawing) of the excitation light and the detection light. In order to make the outer diameter of the optical fiber 103 substantially the same as the outer diameter of the lens 101, a ferrule 104 having the same outer diameter as the outer diameter of the lens 101 is provided so as to surround the optical fiber 103. The optical fiber 103 is fixed by a ferrule 104, and the lens 101 and the ferrule 104 are fixed in a tube 105. Here, the optical fiber 103 and the lens 101 may be in close contact with each other, or a gap may be provided between them.
[0074]
The lens 101 is a gradient index rod lens made of a columnar transparent body whose refractive index continuously changes from the center to the periphery (for example, Japanese Patent Publication No. 63-63502), and is made of, for example, glass or plastic. Is done.
[0075]
In the refractive index distribution type rod lens, the refractive index n (r) at a position at a distance of r in the radial direction from the central axis is represented by n. ₀ , Approximately with quadratic distribution constant g, quadratic equation about r
n (r) = n ₀ ｛1- (g ² / 2) · r ² ｝
Is known as a converging light transmission body represented by
[0076]
The gradient index rod lens has a length z ₀ Is 0 <z ₀ When selected within the range of <π / 2g, the imaging characteristic is the same as that of a normal convex lens while both end surfaces are flat,
s ₀ = Cot (gz ₀ ) / N ₀ g
A focus is created at the location.
[0077]
The gradient index rod lens is manufactured, for example, by the following method.
[0078]
That is, after forming a rod with glass, the glass rod is treated in an ion exchange medium such as potassium nitrate, and ion exchange between monovalent ions in the glass and potassium ions in the ion exchange medium is performed. Inside, a refractive index distribution that continuously decreases from the center to the periphery is provided.
[0079]
Since the bottom surface of the gradient index rod lens is flat, the lens 101 can be easily attached to the end face of the optical fiber 103, and the optical axis of the lens 101 composed of the gradient index rod lens and the optical axis of the optical fiber 103. Can be easily matched. The gradient index rod lens has a cylindrical shape, and the optical fiber 103 connected to the lens 101 can be easily formed into a cylindrical shape. This makes it extremely easy for the jig 102 to hold the optical fiber 103 and the lens 101. Further, since the gradient index rod lens is considerably smaller than the microscope objective lens, the entire system can be miniaturized.
[0080]
The reason why the optical fiber 103 is set to the single mode is that, when detecting a small amount of solute in a sample using photothermal conversion spectroscopy, the excitation light is narrowed as small as possible to increase the energy available for photothermal conversion, and the excitation light is increased. This is because it is desirable that the thermal lens generated by the method be a small lens having small aberration.
[0081]
The excitation light used to generate the thermal lens preferably has a Gaussian distribution. This is because the light emitted from the single mode optical fiber 103 always has a Gaussian distribution, so that the focal point of the excitation light can be reduced.
[0082]
Further, when the thermal lens generated by the excitation light is small, it is desirable to reduce the detection light as small as possible in order to increase the amount of the detection light transmitted through the thermal lens as much as possible. It is also preferable that the laser beam also has a Gaussian distribution like the pump light and propagates in the single mode optical fiber 103.
[0083]
Similarly, when detecting a small amount of solute in a sample using LIF analysis, the higher the energy used for fluorescence emission, the more efficiently the sample emits fluorescence, and the distribution of the sample emitting fluorescence is Since the smaller the size, the more efficiently the fluorescence can be collected, it is desirable that the excitation light for LIF analysis also has a Gaussian distribution capable of narrowing the focus and propagates through the single-mode optical fiber 103.
[0084]
Any type of optical fiber 103 can be used as long as it transmits the excitation light and the detection light. However, when a multi-mode optical fiber is used, the outgoing light does not have a Gaussian distribution, and its bending does not occur. Since the emission pattern changes depending on various conditions such as the condition, stable emission light cannot always be obtained. For this reason, it may be difficult to measure a small amount of solute and the measured value may not be stable. Therefore, as described above, the optical fiber 103 is preferably of a single mode.
[0085]
A light source 106 for excitation light, a light source 107 for detection light, and a two-wavelength multiplexing element 109 are arranged near the end of the optical fiber 103 opposite to the insertion end. The excitation light and the detection light may be made coaxial by using a dichroic mirror or the like without using the two-wavelength multiplexing element 109 and then made incident on the optical fiber 103.
[0086]
The light receiving section 1b for receiving the detection light transmitted through the microchemical system chip 20 in the photothermal conversion spectroscopy is installed at a position facing the lens 101 with the flow path 204 inside the microchemical system chip 20 therebetween. A photoelectric converter 401 for detecting light, a wavelength filter 402 that separates excitation light and detection light and selectively transmits only detection light, and modulates a signal obtained from the photoelectric converter 401 and excitation light. A lock-in amplifier 403 for synchronizing with the modulator 108 used for the synchronization is provided, and a computer 404 for analyzing a signal of the lock-in amplifier 403. Further, in order to selectively transmit only a part of the detection light, the member having the pinhole is positioned such that the pinhole is located on the optical path of the detection light and at a position upstream of the photoelectric converter 401. It may be arranged.
[0087]
In the LIF analysis, the light receiving section 1c (other detection means) for receiving the fluorescence emitted from the sample flowing through the flow path 204 in the microchemical system chip 20 is a fluorescent light concentrator embedded in the microchemical system chip 20. Lens 301, an optical fiber 302 that propagates the fluorescence collected by the condenser lens 301, a detector 303 that detects the fluorescence that has propagated through the optical fiber 302, and blocks light other than the target fluorescence. , A filter 304 used to improve the sensitivity of the LIF analysis.
[0088]
In the LIF analysis, when the excitation light used to excite the fluorescent molecules during the fluorescence measurement enters the detection optical system, it causes noise and lowers the measurement sensitivity. It can be embedded inside the chemical system chip 20 so that the excitation light does not enter the detection optical system so as to be orthogonal to the optical axis of the excitation light. Further, instead of embedding the fluorescence condensing lens 301, it may be attached to the outer surface of the microchemical system chip 20.
[0089]
In the LIF analysis, it is not necessary to modulate the excitation light at the time of measurement, but in the case of modulation, the signal of the detector 303 can be processed by the lock-in amplifier 403 in order to synchronize with the modulation of the excitation light.
[0090]
In order to receive more fluorescent light emitted from the sample flowing through the flow channel 204 in the microchemical system chip 20, it is desirable that the fluorescent light focusing lens 301 be as close to the flow channel 204 as possible.
[0091]
In order to receive as much fluorescence as possible, it is desirable that a condenser lens is provided at the tip of the light receiving section 1c. As described above, the refractive index distribution type rod lens acting as a condenser lens is attached to the tip of the optical fiber 302 embedded in the microchemical system chip 20. Since the gradient index rod lens 301 has a cylindrical shape and a flat end face, it can be easily attached to the tip of the optical fiber 302, and the gradient index rod lens 301 and the optical fiber The optical axis of 302 can be easily adjusted. Further, since the gradient index rod lens 301 is small, it is not necessary to form a large hole for the gradient index rod lens 301.
[0092]
In particular, when a gradient index rod lens 301 having the same diameter as the optical fiber 302 is attached to the end face of the optical fiber 302, the refractive index is formed in a space provided near the flow path 204 in the microchemical system chip 20. The installation is completed only by inserting the optical fiber 302 with the distributed rod lens 301 attached to the tip.
[0093]
It is desirable that a condenser lens is attached to the tip of the optical fiber 302, but LIF analysis is possible even if only the optical fiber 302 is inserted without using a lens. Further, in the case where the gradient index lens 301 is provided, spatial light may be used without using the optical fiber 302.
[0094]
Further, an optical waveguide may be used instead of the optical fiber 302 used for transmitting the fluorescence.
[0095]
As the optical fiber 302 used for the light receiving section 1c, it is desirable to transmit the fluorescence emitted from the sample to the detector 303 as much as possible.
[0096]
According to the first embodiment of the present invention, since photothermal conversion spectroscopy and LIF analysis can be performed only with the microchemical system 1, the range of samples that can be detected is very wide, and a highly versatile device is provided. In addition, since it is not necessary to separately arrange the photothermal conversion spectrometer and the LIF analyzer, the space and cost can be significantly reduced. In addition, since the excitation light used for photothermal conversion spectroscopy and LIF analysis is transmitted from the same light source to the irradiation lens via the optical fiber 103, the microchemical system 1 can be made more compact, and adjustment is not required for each measurement. Thus, the measurement sensitivity of the microchemical system 1 can be improved.
[0097]
Further, since the light receiving section 1b used for photothermal conversion spectroscopy and the light receiving section 1c used for LIF analysis are separately installed, it is possible to perform photothermal conversion spectroscopy and LIF analysis at the same time. For this reason, when a fluorescent substance and a non-fluorescent substance are present in the sample to be measured, they can be detected at the same time, so that it is not necessary to change the apparatus for each of the detection targets, thereby improving the work efficiency of analysis and the like. When the reaction process is followed, a fluorescent substance and a non-fluorescent substance can be simultaneously detected, so that the reaction mechanism can be clarified and the reaction yield can be easily identified. In addition, since a fluorescent substance can be simultaneously detected by the LIF analysis and the photothermal conversion spectroscopy, the correlation between the results of the two can be easily separated from noise, and the measurement sensitivity can be improved.
[0098]
The focal position of the excitation light of the gradient index rod lens 101 needs to be located in the flow path 204 of the microchemical system chip 20. The gradient index rod lens 101 does not need to be in contact with the microchemical system chip 20, but when it is in contact, the thickness of the upper glass plate 201 of the microchemical system chip 20 depends on the refractive index gradient rod lens 101. Adjustable focal length. When the thickness of the upper glass plate 201 is insufficient, a spacer for adjusting the focal length may be provided between the refractive index distribution type rod lens 101 and the upper glass plate 201. If the focal position of the excitation light is fixed in the flow path 204 of the microchemical system chip 20 in this manner, the focal length does not need to be adjusted, and the microchemical system 1 can be further miniaturized.
[0099]
The refractive index distribution type rod lens 101 is set such that the focus position of the detection light is slightly shifted by ΔL from the focus position of the excitation light (FIG. 4A). Ic is the confocal length (nm), and Ic = π · (d / 2) ² / Λ ₁ Is calculated. Here, d is d = 1.22 × λ. ₁ Is the Airy disk calculated by / NA ₁ Is the wavelength (nm) of the excitation light, and NA is the numerical aperture of the gradient index rod lens. In the present embodiment in which the lens 101 is connected to the optical fiber 103, the numerical aperture of the light emitted from the optical fiber is smaller than that of the rod lens, and therefore, it is necessary to use the numerical aperture of the optical fiber 103 for calculating the confocal length. is there.
[0100]
The ΔL value changes depending on the thickness of the sample to be measured. When measuring a sample thinner than the confocal length, the ΔL value is most preferably ΔL = √3 · Ic.
[0101]
Since the value of ΔL represents the difference between the focal position of the detection light and the focal position of the excitation light, the value of ΔL may be a case where the focal length of the detection light is longer than the focal length of the excitation light or a case where it is shorter. The same is true.
[0102]
If the tip of the optical fiber 103 is processed into a spherical shape or the like to form a lens, the excitation light and the detection light can be narrowed without attaching the lens 101 to the tip of the optical fiber 103. Even when the detection light has a wavelength different from the wavelength of the excitation light as in the embodiment, the focal positions of the excitation light and the detection light are substantially the same because there is almost no chromatic aberration of the lens. Therefore, there is a problem that the signal of the thermal lens is hardly detected. Further, since the lens obtained by processing the tip of the optical fiber 103 has a large aberration, there is also a problem that the focus of the excitation light and the detection light is large. Therefore, in the present embodiment, the lens 101 connected to the tip of the optical fiber 103 is made of a lens having chromatic aberration. Accordingly, the focal positions of the excitation light and the detection light can be shifted without using an external optical system, and the microchemical system 1 can be downsized.
[0103]
FIG. 2 is a schematic diagram showing a schematic configuration of a microchemical system according to the second embodiment of the present invention.
[0104]
In FIG. 2, in the microchemical system 2 according to the second embodiment, the same components as those in the microchemical system 1 according to the first embodiment are denoted by the same reference numerals, and description thereof is omitted. The second embodiment is different from the first embodiment in that the light receiving unit 1c is not provided and the light receiving unit 2b also receives light for LIF analysis.
[0105]
Also, when performing photothermal conversion spectroscopy, a filter 402 that blocks light other than the target detection light is required, while when performing LIF analysis, a filter 501 that blocks light other than the target fluorescence is required. is necessary. Therefore, the filter is replaced according to the measurement. Although not shown in FIG. 2, the sensitivity of the LIF analysis can be improved by replacing not only the filter but also the entire detector.
[0106]
In the configuration of FIG. 2, since the light receiving section 2b is located on the optical axis of the excitation light, the excitation light enters the measurement optical system in the LIF analysis. However, if a holographic notch filter or the like that cuts off the excitation light is used as the filter 501 that is replaced when performing the LIF analysis, the LIF analysis can be performed although the sensitivity is lower than in the first embodiment.
[0107]
In the present embodiment, since the light receiving section 2b is shared between the photothermal conversion spectroscopy analysis and the LIF analysis, the system can be made very small as compared with the microchemical system 1 of the first embodiment.
[0108]
【The invention's effect】
As described above in detail, according to the microchemical system according to claim 1, in a microchemical system including an irradiation unit and a detection unit for photothermal conversion spectroscopy, fluorescence emitted from a sample irradiated with excitation light is used. Is provided with another detection means for detecting the chrominance, so that it is possible to provide a microchemical system in which both detection methods of photothermal conversion spectroscopy and laser-induced fluorescence analysis can be used. As a result, for a sample in which a fluorescent substance and a non-fluorescent substance are mixed, the fluorescent substance and the non-fluorescent substance can be detected at the same time, the work efficiency of the user can be improved, and the reaction process and the yield can be identified. For a sample consisting only of a fluorescent substance, photothermal conversion spectroscopy and laser-induced fluorescence analysis can be performed simultaneously, and by correlating the results of the two, separation from noise is facilitated to improve measurement sensitivity. be able to.
[0109]
According to the microchemical system according to the second aspect, since the other detection means is arranged at a position not on the optical axis of the excitation light, even if the photothermal conversion spectroscopy analysis and the laser-induced fluorescence analysis are performed simultaneously, It is possible to prevent excitation light detected as noise from being incident on the detection means, and to prevent a decrease in measurement sensitivity of laser-induced fluorescence analysis. Preferably, the other detecting means is arranged at a position orthogonal to the optical axis of the excitation light.
[0110]
According to the microchemical system according to claim 3, the transparent chemical substrate includes a transparent substrate having a channel for accommodating a sample, and the other detecting means includes a fluorescent light focusing lens, and the fluorescent light focusing lens is provided in the transparent substrate. Embedded in the peripheral portion of the flow path or disposed on the outer surface of the substrate, more fluorescence emitted from the sample flowing through the flow path can be received, and the measurement sensitivity of the laser-induced fluorescence analysis can be improved.
[0111]
According to the microchemical system of the fourth aspect, since one path for transmitting the excitation light and the detection light to the irradiation lens is provided, the excitation light and the detection light are always coaxial, and the optical axis alignment is not required. it can.
[0112]
According to the microchemical system according to the fifth aspect, since one path for propagating the excitation light and the detection light to the irradiation lens is formed of an optical fiber, the excitation light and the detection light can be respectively transmitted without being affected by external changes. It can propagate to the illumination lens.
[0113]
According to the microchemical system according to claim 6, since the optical fiber constituting one path is a single mode fiber, the thermal lens generated by the excitation light becomes a small thermal lens with small aberration, Measurement sensitivity can be improved.
[0114]
According to the microchemical system according to claim 7, the detection light has a wavelength different from the wavelength of the excitation light, and the irradiation lens is formed of a lens having chromatic aberration. It can be shifted without using an optical system, and thus the microchemical system can be downsized.
[0115]
According to the microchemical system of the present invention, since the irradiation lens is a rod lens, it is easy to match the optical axes of the excitation light and the detection light with the optical axis of the rod lens, and the irradiation lens can be downsized. Therefore, the size of the microchemical system can be further reduced.
[0116]
According to the microchemical system according to the ninth aspect, since the irradiation lens is a gradient index rod lens, the size of the irradiation lens can be further reduced, so that the size of the microchemical system can be further reduced.
[0117]
According to the microchemical system of the present invention, since the output modulator for modulating the output of the excitation light is provided, in the photothermal conversion spectroscopy, the thermal lens formed by the irradiation of the excitation light is excited so as not to be saturated. It is not necessary to install a device such as a chopper for periodically turning on and off the light irradiation, and the transmission path of the excitation light can be entirely within the optical fiber, miniaturizing the microchemical system. In addition, the excitation light can be propagated to the irradiation lens without being affected by external changes, so that the measurement sensitivity of the microchemical system can be improved.
[0118]
According to the microchemical system according to claim 11, in the microchemical system including the irradiation unit and the detection unit for photothermal conversion spectroscopy, the detection unit also detects fluorescence emitted from the sample by irradiation with the excitation light, The whole system can be made compact.
[0119]
According to the microchemical system according to the twelfth aspect, the detection light transmitting unit that transmits only the detection light and the fluorescence transmission unit that transmits only the fluorescence can be moved in and out between the detection unit and the sample on the optical axis of the excitation light. Since it is installed, when performing photothermal conversion spectroscopy, the detection light transmission means is inserted on the optical axis of the excitation light, and when performing LIF analysis, the fluorescence transmission means is inserted on the optical axis of the excitation light. The analysis contents can be switched.
[0120]
According to the microchemical system according to the thirteenth aspect, since one path for transmitting the excitation light and the detection light to the irradiation lens is provided, the excitation light and the detection light are always coaxial, and the optical axis alignment is not required. it can.
[0121]
According to the microchemical system according to claim 14, one path for transmitting the excitation light and the detection light to the irradiation lens is formed of an optical fiber, so that the excitation light and the detection light are not affected by external changes. It can propagate to the illumination lens.
[0122]
According to the microchemical system of claim 15, since the optical fiber constituting one path is a single mode fiber, the thermal lens generated by the excitation light becomes a small thermal lens with small aberration, Measurement sensitivity can be improved.
[0123]
According to the microchemical system according to claim 16, the detection light has a wavelength different from the wavelength of the excitation light, and the irradiation lens is formed of a lens having chromatic aberration. It can be shifted without using an optical system, and thus the microchemical system can be downsized.
[0124]
According to the microchemical system of the present invention, since the irradiation lens is a rod lens, it is easy to align the optical axis of the excitation light and the detection light with the optical axis of the rod lens, and the irradiation lens can be downsized. Thus, the size of the microchemical system can be further reduced.
[0125]
According to the microchemical system of the present invention, since the irradiation lens is a gradient index rod lens, the size of the irradiation lens can be further reduced, so that the size of the microchemical system can be further reduced.
[0126]
According to the microchemical system according to the nineteenth aspect, since the output chemical modulator for modulating the output of the excitation light is provided, in the photothermal conversion spectroscopy, the thermal lens formed by the irradiation of the excitation light is excited so as not to be saturated. It is not necessary to install a device such as a chopper for periodically turning on and off the light irradiation, and the transmission path of the excitation light can be entirely within the optical fiber, miniaturizing the microchemical system. In addition, the excitation light can be propagated to the irradiation lens without being affected by external changes, so that the measurement sensitivity of the microchemical system can be improved.
[Brief description of the drawings]
FIG. 1 is a schematic diagram showing a schematic configuration of a microchemical system according to a first embodiment of the present invention.
FIG. 2 is a schematic diagram illustrating a schematic configuration of a microchemical system according to a second embodiment of the present invention.
FIG. 3 is an explanatory view of the principle of a thermal lens.
4A and 4B are explanatory diagrams of a formation position of a thermal lens with respect to an optical axis (Z-axis direction) of excitation light and a focus position of detection light, wherein FIG. 4A shows a case where the objective lens has chromatic aberration, Indicates a case where the objective lens has no chromatic aberration.
FIGS. 5A and 5B are explanatory diagrams of a formation position of a thermal lens with respect to an optical axis of the excitation light (Z-axis direction) and a focus position of the detection light; FIG. (B) shows the case where the thermal lens is formed farther than the focal position of the detection light.
FIG. 6 is an explanatory diagram of a method for detecting a change in the refractive index of a thermal lens in a conventional photothermal conversion analyzer. Also shows a case where the focal position is set to be far away.
[Explanation of symbols]
1,2 Micro chemical system
1a Irradiation unit
1b, 1c, 2b Light receiving unit
20 Micro Chemical System Chips
101 lens
103 Optical fiber
106 Light source for excitation light
107 Light source for detection light
204 channel
301 Fluorescence focusing lens
302 Optical Fiber
303,401 photoelectric converter
304, 402, 501 filters
403 lock-in amplifier

Claims (19)

Irradiation means for irradiating the sample with excitation light and detection light for photothermal conversion spectroscopy via an irradiation lens, and detection for detecting the detection light transmitted through the thermal lens generated in the sample by irradiation of the excitation light A microchemical system comprising:
A microchemical system comprising: another detection unit configured to detect fluorescence emitted from the sample by irradiation with the excitation light. 2. The microchemical system according to claim 1, wherein the other detection unit is arranged at a position that is not on an optical axis of the excitation light. A transparent substrate having a flow path for accommodating the sample, wherein the other detection means includes a fluorescent light focusing lens, wherein the fluorescent light focusing lens is embedded in a peripheral portion of the flow path in the transparent substrate or an outer surface of the substrate. The microchemical system according to claim 2, wherein the microchemical system is disposed in a microchemical system. The microchemical system according to any one of claims 1 to 3, further comprising one path for propagating the excitation light and the detection light to the irradiation lens. 5. The microchemical system according to claim 4, wherein said one path comprises an optical fiber. The microchemical system according to claim 5, wherein the optical fiber is a single mode fiber. The microchemical system according to any one of claims 1 to 6, wherein the detection light has a wavelength different from the wavelength of the excitation light, and the irradiation lens includes a lens having chromatic aberration. The microchemical system according to claim 7, wherein the irradiation lens is a rod lens. 9. The microchemical system according to claim 8, wherein the irradiation lens is a gradient index rod lens. The microchemical system according to any one of claims 1 to 9, further comprising an excitation light output adjusting device that modulates an output of the excitation light. Irradiation means for irradiating the sample with excitation light and detection light for photothermal conversion spectroscopy via an irradiation lens, and detection for detecting the detection light transmitted through the thermal lens generated in the sample by irradiation of the excitation light A microchemical system comprising:
The said detection means also detects the fluorescence emitted from the said sample by irradiation of the said excitation light, The microchemical system characterized by the above-mentioned. The microchemical system further includes a detection light transmission unit that transmits only the detection light and a fluorescence transmission unit that transmits only the fluorescence,
12. The microchemical system according to claim 11, wherein the detection light transmission unit and the fluorescence transmission unit are installed on the optical axis of the excitation light so as to be able to be inserted and removed between the detection unit and the sample. The microchemical system according to claim 12, further comprising one path for propagating the excitation light and the detection light to the irradiation lens. 14. The microchemical system according to claim 13, wherein said one path comprises an optical fiber. The microchemical system according to claim 14, wherein the optical fiber is a single mode fiber. The microchemical system according to any one of claims 11 to 15, wherein the detection light has a wavelength different from a wavelength of the excitation light, and the irradiation lens is formed of a lens having chromatic aberration. 17. The microchemical system according to claim 16, wherein the irradiation lens is a rod lens. 18. The microchemical system according to claim 17, wherein the irradiation lens is a gradient index rod lens. The microchemical system according to any one of claims 11 to 18, further comprising an excitation light output adjusting device that modulates an output of the excitation light.

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