JP4357399B2 - Electrophoresis device - Google Patents

Description

  The present invention relates to a technique for separating and analyzing nucleic acids, proteins and the like by electrophoresis, for example, a capillary electrophoresis apparatus.

  For example, a capillary electrophoresis apparatus as disclosed in JP-A-10-19846 uses a quartz tube and a capillary made of a polymer covering the same for the purpose of determining the DNA base sequence and base length. Electrophoretic apparatus. A sample containing DNA to be measured is introduced into a separation medium such as polyacrylamide packed in a quartz capillary, and a voltage is applied to both ends of the capillary. The DNA synthesis product in the sample is electrophoresed in the capillary and is separated according to the size of the molecular weight or the like to generate a DNA band in the capillary. A fluorescent dye is added to each DNA band, which develops color when irradiated with laser light, which is read by a fluorescence measuring means to determine the DNA sequence. Protein separation and analysis can be performed in the same manner to examine the structure of the protein.

  In Japanese Patent Application Laid-Open No. 10-19846, fluorescence caused by laser light irradiation is propagated into the capillary, taken out from the end face of the capillary, and detected by a photodetector. In addition to the above publications, USP 5,730,850, USP 6,661,510, and USP 6,613,212 disclose an electrophoresis apparatus that forms an image of the end face of a capillary on a detector.

JP-A-10-19846 USP 5,730,850 USP 6,661,510 USP 6,613,212

  As a result of intensive investigations by the inventor of the present invention, an inventor of the present application has found the following problem with respect to an end detection method in which the end surface of the capillary arranged two-dimensionally is imaged on the light receiving surface of a two-dimensional detector such as a CCD .

  When imaging the end face of the capillary on the light receiving surface, if the position of the light receiving surface is adjusted so that all the end faces of the capillary arranged in two dimensions are in focus, several wavelengths are emitted from one capillary end. The component is out of focus due to chromatic aberration of the camera lens. That is, the present inventors have found a problem that if the position of the two-dimensional detector is set on the imaging surface of a specific wavelength, the focus is not achieved at other wavelengths, that is, the image becomes out of focus.

  An object of the present invention relates to detecting a plurality of wavelength components emitted from a capillary end face with high sensitivity.

  The present invention relates to the provision of a chromatic aberration correction mechanism in an electrophoretic apparatus that detects a plurality of wavelengths emitted from a capillary end face arranged in two dimensions by a two-dimensional detector. For example, a plane formed by a plurality of capillary ends is inclined with respect to the optical axis of the condenser lens, and an imaging plane having a plurality of wavelengths is on the same plane. In addition, for example, multiple wavelength light emitted from the end face of the capillary is transmitted through transparent members having different thicknesses corresponding to the multiple wavelengths, and the imaging surfaces of the multiple wavelengths are on the same plane.

  According to the present invention, the influence of chromatic aberration can be reduced, photodetection sensitivity can be increased, and crosstalk can be reduced.

  The above and other novel features and benefits of the present invention will be described below with reference to the drawings. However, the drawings are for explanation only, and do not limit the scope of the present invention.

  FIG. 1 shows a schematic perspective view of the electrophoresis apparatus. FIG. 2 is a schematic perspective view of the capillary head. FIG. 3 shows a schematic sectional view in the vicinity of the polymer filling block. FIG. 4 shows a schematic flow diagram of the analysis procedure. The outline of the electrophoresis apparatus will be described below with reference to these drawings.

  The electrophoresis apparatus in the present embodiment (hereinafter referred to as “this apparatus”) uses a plurality of capillaries filled with an electrophoresis medium, introduces a sample into each capillary, and separates and analyzes the sample components in the sample. To do. For example, a base sequence can be analyzed by simultaneously analyzing a plurality of samples containing DNA. The basic configuration of this apparatus is a capillary array, an irradiation optical unit, a detection optical unit, an autosampler unit, an electrophoresis medium filling unit, a power supply unit, and a temperature control unit.

  The capillary array 101 is a detachable exchange member composed of a plurality of capillaries 102, and is exchanged for a new one when the quality deteriorates due to a predetermined number of analyzes and the separation performance decreases. When the measurement technique is changed, the length of the capillaries 102 can be adjusted by exchanging the capillaries 102 with capillaries 102 having different lengths. The capillary array 101 includes a sample introduction unit 104 that introduces a sample into the capillary 102, an irradiation unit 103 that irradiates the separated sample with excitation light, and a capillary head 107 that bundles a plurality of capillaries.

  The capillary 102 is a thin tube having an inner diameter of several tens to several hundreds of microns and an outer diameter of several hundreds of microns, and the surface is coated to improve the strength. The electrophoresis path is formed by filling the inside of the capillary 102 with the electrophoresis medium. By applying a voltage to both ends of the electrophoresis path, the sample can be electrophoretically separated.

  In this embodiment, a quartz pipe whose outer surface is coated with fluorine is used so that light generated in the capillary 102 propagates through the inside. In the irradiation unit 103 slightly away from the sample introduction unit 104, the fluorine coating is removed in order to efficiently irradiate the sample in the capillary 102 with the laser beam 112. A plurality of capillaries 102 are bundled to form a capillary array 101. The number of capillaries 102 is, for example, 384, 192, 96, and 8. If necessary, the capillary 102 may be coated with polyimide.

  The sample introduction unit 104 has the sample introduction end 105 of the capillary 102 arranged in alignment with the well of the sample container 147, and can introduce a plurality of samples held in each well into the migration path.

  In this embodiment, the sample introduction end 105 is formed by inserting the tip of the capillary 102 into the hollow electrode 106, and the sample introduction end 105 is arranged in 24 rows and 16 columns to constitute the sample introduction portion 104. At the sample introduction end 105, the tip of the capillary 102 protrudes slightly from the hollow electrode 106. The hollow electrode 106 composed of a stainless steel pipe is electrically connected to a high voltage power source 162. By immersing the sample introduction end 105 in the sample and applying a voltage, the sample can be electrophoresed and introduced into the capillary. The sample introduction method is not limited to electrophoresis, and the sample may be introduced into the electrophoresis path by pressure or dispensing.

  The irradiation unit 103 is a part that irradiates the electrophoresis medium filled in the capillary 102 with the laser light 122 that is excitation light, and can irradiate the sample component subjected to electrophoretic separation with excitation light. The phosphor labeled on the sample component emits light having a wavelength depending on the sample component.

  In the present embodiment, a plurality of capillaries 102 are divided into a plurality of groups, and a plurality of capillaries 102 are arranged on a plurality of glass substrates to form several capillary arrays. In addition, the number of sets of capillary arrays is one set, two sets, three sets, four sets, and the like. On each glass substrate, the capillaries 102 are arranged substantially parallel to each other and substantially parallel to the glass substrate. In addition, the removed portions of the fluorine film in each capillary 102 are arranged in a straight line. Here, the fact that the capillaries 102 are substantially parallel to each other and parallel to the glass substrate means that the degree of parallelism is within a precision error. The plurality of laser beams 122 irradiated from both sides of each glass substrate by the irradiation optical unit propagate through the polyimide film removal portions in the plurality of capillaries 102, respectively, and excite the sample in each capillary 102 at the same time.

  The capillary head 107 holds a plurality of capillaries in a bundle and can be attached to and detached from the apparatus main body. As shown in FIG. 2, the capillary head 107 of the present embodiment holds the end portions 108 of the capillaries so as to form a matrix. A part of the fluorescence generated from the sample component in the irradiation unit 103 is totally reflected on the inner surface of the capillary 102 and propagates through the capillary 102, is emitted from the terminal unit 108, and is detected by the detection optical unit. Further, the capillary head 107 can be connected to the polymer filling block 152 in a pressure-resistant and airtight manner. Then, a new electrophoresis medium can be filled into the capillary 102 from the end portion 108 by the electrophoresis medium filling unit. Note that the termination units 108 may be arranged in a matrix of 96 rows and 4 columns, 64 rows and 6 columns, or 48 rows and 8 columns as needed, or may be simply converged into one.

  The irradiation optical unit is a mechanism that irradiates the irradiation unit 103 of the capillary array 101 with laser light 122 that is excitation light. The irradiation optical unit of this embodiment includes a laser light source 112, a beam splitter A124, a mirror A125, a beam splitter B126, a mirror B127, and a condenser lens 128.

  The laser light source 112 can oscillate laser light that is excitation light for exciting sample components. The beam splitter A124 and the mirror A125 divide the laser beam 122 into two parts so that the capillary array of the irradiation unit 103 can be irradiated from both sides. The plurality of beam splitters B126, the plurality of mirrors B127, and the condenser lens 128 respectively adjust the plurality of laser beams 122 and irradiate the light irradiation unit 103 with the laser beams 122. Laser light 122 applied to each capillary array propagates to adjacent capillaries 102 one after another.

As the laser light source 112, a liquid laser, a gas laser, or a semiconductor laser can be used as appropriate, and an LED may be substituted if necessary. For example, an argon ion laser is a multi-line that oscillates a laser 122 with wavelengths of 488 nm and 514.5 nm and an output of 100 mW. In addition to a multiline, a single line may be used, or two or more types of laser light sources may be combined. Further, the wavelength of the laser beam 122 may be 532 nm or 633 nm as appropriate. Further, the laser beam 122 may be irradiated only from one side of the capillary array, or the laser beam 122 may be irradiated in a time division manner.

  The detection optical unit 131 is a mechanism for detecting light emitted from the sample component. As shown in FIG. 3, the detection optical unit 131 of this embodiment includes a first camera lens 118, an optical prism 134, a second camera lens 135, and a two-dimensional detector 136.

  The end face of the capillary head 107 having the end portion 108 is filled with the electrophoresis medium and disposed so as to face the detection window 132 made of non-fluorescent quartz glass through the polymer flow path 154. The light emitted from the end portion 108 passes through the migration medium in the polymer flow path 154, is radiated from the detection window 132, enters the first camera lens 133, and becomes a parallel light flux. The parallel light flux is wavelength-dispersed by the optical prism 134 and imaged on the two-dimensional detector 136 which is a CCD camera by the second camera lens 135. Then, the signal from the CCD camera is processed by a computer to analyze the sample.

  Instead of the optical prism 134, the wavelength dispersion means may be configured by appropriately combining diffraction gratings and filters. The two-dimensional detector 136 may be configured by appropriately combining a one-dimensional detector, a photomultiplier, a photodiode, and the like and an optical mechanism.

The autosampler unit is a mechanism that transports and holds each container used for electrophoretic analysis including the sample container 147 to a predetermined position such as directly below the sample introduction unit 104. The autosampler unit of the present embodiment transports the sample container 147, the buffer container 144, the cleaning container 145, and the waste liquid container 146 by the robot arm 142 having the claws 143.
The robot arm 142 includes a claw 143 for fixing each container thereon, and can move three-dimensionally. As a result, each container stored in a predetermined place can be transported directly below the sample introduction unit 104, held at that place for a predetermined time, and returned to the predetermined place.

The buffer container 144 is a container that holds a buffer solution that immerses the sample introduction end 105. During the migration analysis, the buffer solution 144 is transported directly below the sample introduction unit 104 so that the buffer solution immerses the sample introduction end 105. Further, during the standby state of the apparatus, the sample is transported in the same manner as in the electrophoresis analysis, and the sample introduction end 105 is immersed in a buffer solution to prevent the electrophoresis medium in the capillary 102 from being dried.

  The cleaning container 145 is a container that holds a cleaning liquid for cleaning the sample introduction end 105, and is transported directly below the sample introduction unit 104 after loading of the electrophoresis medium, preliminary migration, and sample introduction. By immersing the sample introduction end 105 in the cleaning liquid in the cleaning container, the sample introduction end 105 can be cleaned and contamination can be avoided.

  The waste liquid container 146 is a container for holding a used electrophoresis medium, and receives the used electrophoresis medium that is transported directly below the sample introduction unit 104 when filling the electrophoresis medium and discharged from the sample introduction end 105 when filling the electrophoresis medium. .

The sample container 147 is a container that holds a plurality of trace samples, and is transported immediately below the sample introduction unit 104 when the sample is introduced. In this embodiment, a sample container 147 is configured by placing a septa, which is a resin sheet, on a sample plate having 24 rows and 16 columns of wells capable of holding a sample of several tens of μl, and sandwiching it with a holder and a clip. ing. The sample is, for example, a solution that is fluorescently labeled so as to identify four types of nucleoside base molecules and contains a large number of nucleic acids having an appropriate length (size). The septa usually have a sealed through hole at a position corresponding to the well, and the sample introduction end 105 and the sample can be contacted at the time of sample introduction while preventing the sample in the well from evaporating. Also, a protective film can be attached to the upper surface of the septa to prevent sample evaporation. In addition, the holder and the clip sandwich a sample plate and a septa between them, and are integrated to form a sample container 147 that can be transported by the robot arm 142.

  The electrophoresis medium filling unit shown in FIGS. 1 and 3 is a mechanism for filling the capillary 102 with a polymer as the electrophoresis medium. The electrophoresis medium filling unit of this embodiment includes a polymer filling block 152, a syringe 153, a tube 155, and an electromagnetic valve 156, and can automatically fill the capillary 102 with a new electrophoresis medium before starting analysis.

  The polymer filling block 152 has a polymer flow path 154 and a detection window 132, is connected to the syringe 153 and the tube 155, and the capillary head 110 can be attached and detached. The capillary head 110 is mounted on the polymer filling block 152 while maintaining pressure and air tightness. The terminal portion 108 of the capillary head 107 is disposed so as to face the detection window 132, and a region sandwiched between the terminal portion 108 and the detection window 132 forms a part of the polymer flow path 154. The polymer flow path 154 communicates with a syringe 153 filled with a migration medium and a tube 155 provided with an electromagnetic valve 156. The other end of the tube 155 is immersed in a buffer solution held in the anode buffer container 163. When filling the capillary 102 with the electrophoresis medium, the waste liquid container 146 is disposed immediately below the sample introduction unit 104, the electromagnetic valve 156 is closed, and the plunger of the syringe 153 is pushed. As a result, the electrophoresis medium in the syringe 153 flows into the capillary 102 from the terminal end portion 108 via the polymer channel 154. Further, the used electrophoresis medium in the capillary 102 is discharged from the sample introduction end 105 and received by the waste liquid container 146.

  The power supply unit is a mechanism that applies a voltage to the migration path composed of the migration medium in the capillary 102, and can electrophorese the sample. The power supply unit of this embodiment includes a high voltage power supply 162 that is electrically connected to the hollow electrode 106 and the anode electrode 164 and can generate a high voltage of around 15 kV.

  At the time of sample introduction, the inside of the capillary 102, the polymer channel 154 and the tube 155 is filled with the electrophoresis medium, the sample introduction end 105 is immersed in the sample held in the well of the sample container 147, and the electromagnetic valve 156 is opened. As a result, an energization path comprising the hollow electrode 106, the sample in the well, the capillary 102, the polymer flow path 154, the tube 155, the buffer solution in the anode buffer container 163, and the anode electrode 164 is formed. Then, a pulse voltage is applied to this energization path with the hollow electrode 106 as a negative potential and the anode electrode 164 as a positive potential. As a result, a negatively charged sample component, such as DNA, present in the well is introduced from the sample introduction end 105 into the migration path.

  Also, during the electrophoresis analysis, unlike the sample introduction, the sample introduction end 105 is immersed in a buffer solution held in the buffer container 144. As a result, a current path including the hollow electrode 106, the buffer solution in the buffer solution 144, the capillary 102, the polymer channel 154, the tube 155, the buffer solution in the anode buffer container 163, and the anode electrode 164 is formed. Then, unlike the sample introduction, a high voltage of around 15 kV is applied. As a result, an electric field is generated in the direction from the irradiation unit 103 to the sample introduction unit 104, and the negatively charged sample component introduced into the migration path is electrophoresed in the direction of the irradiation unit 103.

  The temperature control unit is a mechanism for controlling the temperature of the migration path that affects the migration speed of the sample components. The temperature control unit in the present embodiment houses the capillary 102 in a thermostat (not shown). Then, the air maintained at a constant temperature by a temperature control mechanism such as Peltier is circulated in the thermostatic chamber by a blowing mechanism such as a fan, and the capillary 102 is maintained at a predetermined temperature.

  Hereinafter, the basic procedure of electrophoretic analysis will be described mainly with reference to FIG.

  The basic procedure of electrophoretic analysis can be broadly divided into pre-preparation 201, electrophoresis medium filling 203, preliminary electrophoresis 204, sample introduction 205, and electrophoresis analysis 206. The operator of this apparatus sets a sample and a reagent in this apparatus as a preliminary preparation before starting analysis (201). More specifically, first, the buffer container 144 and the anode buffer container 163 are filled with a buffer solution that forms part of the current path. The buffer solution is, for example, an electrolyte solution that is commercially available for electrophoresis from various companies.

  In addition, a sample to be analyzed is dispensed into the well of the sample container 147. The sample is, for example, a DNA PCR product. In addition, a cleaning solution for cleaning the sample introduction unit 104 is dispensed into the cleaning container 145. The cleaning solution is pure water, for example. Further, a separation medium for electrophoresis of the sample is injected into the syringe 153. The electrophoresis medium is, for example, a polyacrylamide separation gel commercially available for electrophoresis from various companies. Furthermore, when the deterioration of the capillary 102 is expected or when the length of the capillary 102 is changed, the capillary array 101 is replaced. Then, after the preliminary preparation is completed, the operator operates the apparatus and starts analysis (202). The electrophoresis medium filling 203 is a procedure for filling the capillary 102 with a new electrophoresis medium and forming an electrophoresis path.

  In the loading of the loading medium 203 in the present embodiment, first, the waste solution 146 is carried directly under the sample introduction unit 104 by the autosampler unit so that the used migration medium discharged from the sample introduction end 105 can be received (211). ). Then, the syringe 153 is driven to fill the capillary 102 with a new electrophoresis medium, and the used electrophoresis medium is discarded (212). Finally, the sample introduction end 105 is immersed in the cleaning solution in the cleaning container 145, and the sample introduction end 105 soiled with the electrophoresis medium is washed (213). The preliminary electrophoresis 204 is a procedure for applying a predetermined voltage to the electrophoresis medium so that the electrophoresis medium is in a state suitable for electrophoresis. In the preliminary electrophoresis 204 in the present embodiment, first, the sample introduction end 105 is immersed in the buffer solution in the buffer container 144 by the autosampler unit to form a current path (214). Then, a voltage of about several to several tens of kilovolts is applied to the electrophoresis medium for several to several tens of minutes by the power supply unit to bring the electrophoresis medium into a state suitable for electrophoresis (215). Finally, the sample introduction end 105 is immersed in the washing solution in the washing container 145, and the sample introduction end 105 soiled with the buffer solution is washed (216). The sample introduction 205 is a procedure for introducing sample components into the migration path.

  In the sample introduction 205 in the present embodiment, first, the sample introduction end 105 is immersed in the sample held in the well of the sample container 147 by the autosampler unit. As a result, an energization path is formed, and the sample component is introduced into the migration path (217). Then, a pulse voltage is applied to the energization path by the power supply unit, and the sample component is introduced into the migration path (218). Finally, the sample introduction end 105 is immersed in the cleaning solution in the cleaning container 145, and the sample introduction end 105 contaminated with the sample is washed (219). The migration analysis 206 is a procedure for separating and analyzing each sample component contained in a sample by electrophoresis. In the electrophoretic analysis 206 in the present embodiment, first, the sample introduction end 105 is immersed in the buffer solution in the buffer container 144 by the autosampler unit to form a current path (220).

  Then, a high voltage of about 15 kV is applied to the energization path by the power supply unit to generate an electric field in the migration path (221). Due to the generated electric field, each sample component in the migration path moves to the irradiation unit 103 at a speed depending on the property of each sample component. That is, the sample components are separated by the difference in moving speed. And it detects in order from the sample component which reached the irradiation part 103. For example, when the sample includes a large number of DNAs having different base lengths, a difference occurs in the moving speed depending on the base length, and the irradiation unit 103 is reached in order from the DNA having the shorter base length. If a fluorescent substance depending on the terminal base sequence is attached to each DNA, the terminal base sequence can be detected in the order of arrival at the irradiation unit 103.

When the planned data is taken, the voltage application is stopped and the electrophoresis analysis is finished (213).

  The above is a series of analysis procedures. When further analysis is performed, the analysis procedure is advanced from the loading of the migration medium 203.

  Hereinafter, a chromatic aberration correction mechanism, which is a characteristic mechanism of the present embodiment, will be described with reference to FIGS.

  We have found that the problem shown in FIG. 5 occurs in the end detection method. FIG. 5 shows a detection system for light emission from the capillary array in the end detection. Light emitted from the end face 501 of the capillary is converted into parallel light by the fluorescence condensing lens 502, dispersed by the prism 503, and imaged on the CCD light receiving surface 505 by the imaging lens 504. The CCD light receiving surface 505 is perpendicular to the imaging lens. On the end face 501 of the capillary, the capillaries are arranged in a 64 × 6 lattice pattern. FIG. 6 shows an image formed on the CCD light receiving surface 505. As shown in FIG. 6, the image 601 in which the light emission from one capillary is dispersed is arranged in the Y-axis direction with 64 and the X-axis direction with 6 images. As shown in FIG. 6, the spectrally emitted light from the 384 capillaries can be detected simultaneously.

Here, the capillary end face 501 is set perpendicularly to the optical axis 506 of the fluorescent condenser lens 502. At this time, as shown in FIG. 5, due to the chromatic aberration of the lens, the surfaces 509 and 510 formed by the image formation points of the dispersed light emitted from the two capillaries 507 and 508 arranged in the X-axis direction are not coplanar. The inventors have found that even if the CCD image forming surface is moved, there arises a problem that not all the images of the dispersed light can be formed on the CCD light receiving surface 505. A part of the light emission becomes out of focus, which causes a problem of reducing the sensitivity of fluorescence detection and increasing crosstalk. It is assumed that an angle formed between the surfaces 509 and 510 formed by the imaging points of the dispersed light and the CCD light receiving surface 505 perpendicular to the optical axis 511 of the imaging lens is θ512.

  Therefore, the inventors made the angle θ703 formed between the end face 701 of the capillary array and the surface perpendicular to the optical axis 702 of the condenser lens the same as the angle θ512 shown in FIG. Here, the capillary axis 705 is parallel to the optical axis 702 of the condenser lens.

  The image plane in this case is as shown in FIG. In other words, the imaging planes 704 coincide with each other for each wavelength of light separated from the plurality of capillaries arranged in the X-axis direction. As described above, in this embodiment, by forming the imaging surface 704 as the light receiving surface of the CCD, the influence of chromatic aberration can be eliminated for all the capillaries and the imaging surface can be on the same plane. It is possible to suppress the increase in talk and realize high sensitivity and low crosstalk of fluorescence detection.

  With reference to FIG. 8, the chromatic aberration correction mechanism which is a characteristic mechanism of the present embodiment will be described.

  As in the first embodiment, the end face 801 of the capillary array is shifted from the optical axis 802 of the lens by θ in FIG. 5 as shown in FIG. However, in the second embodiment, unlike the first embodiment, the capillary axis 803 is perpendicular to the capillary array end face 801 and is not parallel to the condenser lens optical axis 802. The image plane 804 in this case is as shown in FIG. That is, the imaging planes 804 coincide with each other for each wavelength of light separated from the plurality of capillaries arranged in the X-axis direction. As described above, in this embodiment, by using the imaging surface 804 as the light receiving surface of the CCD, the influence of chromatic aberration can be eliminated for all the capillaries, and the imaging surface 804 can be on the same surface. Increase in crosstalk can be suppressed, and high sensitivity and low crosstalk in fluorescence detection can be realized. However, since the fluorescence emission intensity is distributed around the capillary axis 803 and is strongest in the direction of the capillary axis 803, the light is condensed under the conditions of Example 2 in which the capillary axis 803 and the optical axis 802 of the condenser lens do not coincide. The light collection efficiency of the lens 805 is lowered, and the sensitivity of fluorescence detection is inferior to that of the first embodiment.

  With reference to FIG. 9, the chromatic aberration correction mechanism which is a characteristic mechanism of the present embodiment will be described.

In the third embodiment, the configuration / arrangement of the end 901 of the capillary array is the same as in FIG. A wedge-shaped transparent glass plate 907 shown in FIG. 9 is installed in front of the CCD light receiving surface 902. The glass thickness on the red side, which is the long wavelength side, is reduced, and the glass thickness on the blue side, which is the short wavelength side, is increased. As a result, the dispersed fluorescence 905 and 906 can be imaged on the CCD light receiving surface 902 perpendicular to the optical axis 904 of the imaging lens 903 with corrected chromatic aberration. One wedge-shaped transparent glass plate 907 corrects chromatic aberration for 64 capillaries in a row. Six wedge-shaped glass plates 907 may be installed in front of the CCD light-receiving surface 902 in accordance with six rows of capillary imaging positions, or one glass plate connected with six wedge-shaped glasses is installed. May be.

  With reference to FIGS. 10 to 13, a chromatic aberration correction mechanism that is a characteristic mechanism of the present embodiment will be described.

  In Example 4, the configuration and arrangement of the terminal end 1000 of the capillary array are the same as those in FIG. As shown in FIG. 10, four band pass filter groups 1001 that rotate without installing a prism are installed. Thus, the four band pass filters can be switched as appropriate. The four bandpass filters (FIG. 11) transmit only the light of the red transmission bandpass filter 1001, the yellow transmission bandpass filter 1002, the green transmission bandpass filter 1003, and the blue transmission bandpass filter 1004, respectively. Also, four imaging position correction glass plate groups 1010 having different thicknesses for correcting chromatic aberration corresponding to the respective wavelengths of the band pass filter group 1001 are set at positions corresponding to the respective band pass filters. When the pass filter group 1001 rotates, the correction glass plate group 1010 also rotates at the same time (FIG. 11). Only light that passes through the bandpass filter group 1001 is imaged on the CCD 1020 at a position corresponding to each capillary. In the CCD 1020, the charge of the CCD is moved in synchronization with the timing of switching the filters of the bandpass filter group 1001 (FIG. 12). Thereby, an optical signal corresponding to each bandpass filter can be imaged at different positions of the CCD 1020. In the present embodiment, it is possible to form an image on the CCD that is substantially equivalent to that of the first embodiment without using a prism (FIG. 13).

  Since the chromatic aberration is corrected by the correction glass plate group 1010, the fluorescence from the capillary array end 1000 can be imaged on the CCD without defocusing at all wavelengths.

With reference to FIGS. 14 and 15, the chromatic aberration correction mechanism which is a characteristic mechanism of the present embodiment will be described. The configuration of the fluorescence detection type of the fifth embodiment is the same as that of the first embodiment. In the first embodiment, the configuration of the laser irradiation unit is such that a laser beam is irradiated to one or both ends of a capillary array composed of a plurality of capillaries arranged on a flat substrate, and the laser beam is successively applied to adjacent capillaries. In this embodiment, the capillary 1402 is arranged on the same plane 1401 (FIG. 14), and the laser beam 1403 is spread and widened by the cylindrical concave lens 1406 in the capillary array direction in this embodiment. A laser beam 1404 collectively irradiates the same planar capillary 1402 (FIG. 15), and in the capillary axis direction, the laser beam is condensed on the capillary by a cylindrical convex lens 1405 (FIG. 16). Regardless of the laser irradiation method, on the CCD image formation surface, as in the first embodiment, the image formation surfaces of the capillaries in the X-axis direction coincide with each other, and at the same time, the image formation surfaces of the light beams having the respective wavelengths separated from each other. To match. In this way, in this embodiment, the influence of chromatic aberration can be eliminated for all capillaries, and the image plane can be made on the same plane. Therefore, low sensitivity due to defocusing and increase in crosstalk are suppressed, and high sensitivity of fluorescence detection is achieved. And crosstalk reduction can be realized.

  The example of the present invention has been described above, but the present invention is not limited to the above-described example, and various modifications can be made by those skilled in the art within the scope of the invention described in the claims. Will be understood. For example, the embodiments can be appropriately combined.

  Further, light emission from the sample is not limited to four wavelengths, and each embodiment can be appropriately changed according to the case of two wavelengths or three wavelengths.

The schematic perspective view of an electrophoresis apparatus. The schematic perspective view of a capillary head. The schematic sectional drawing of the polymer filling block vicinity. Schematic flow chart of analysis procedure. The layout of the detection system of the end detection method. Image on CCD of end detection method. FIG. 3 is a layout diagram of a detection system of the end detection method according to the first embodiment. FIG. 6 is a layout diagram of a detection system of the end detection method according to the second embodiment. FIG. 9 is a layout diagram of a detection system of the end detection method of Example 3. FIG. 6 is a layout diagram of a detection system of a terminal detection method according to a fourth embodiment. 4 is a bandpass filter / chromatic aberration correcting rotary plate of Example 4. FIG. Image on CCD of end detection method of Example 4. The whole image on CCD of the terminal detection method of Example 4. FIG. FIG. 10 shows a laser beam irradiation part in the capillary array of Example 5. FIG. FIG. 10 is a side view of laser light irradiation in the capillary array of Example 5. FIG. 10 is a top view of laser light irradiation in the capillary array of Example 5.

Explanation of symbols

DESCRIPTION OF SYMBOLS 101 ... Capillary array, 102, 1402 ... Capillary, 103 ... Irradiation part, 104 ... Sample introduction part, 105 ... Sample introduction end, 106 ... Hollow electrode, 107 ... Capillary head,
DESCRIPTION OF SYMBOLS 108 ... Termination part, 121 ... Irradiation optical unit, 122,1403 ... Laser light, 112 ... Laser light source, 124 ... Beam splitter A, 125 ... Mirror A, 126 ... Beam splitter B, 127 ... Mirror B, 131 ... Detection optical unit , 132 ... detection window, 133 ... first camera lens, 134 ... optical prism, 135 ... second camera lens, 136 ... two-dimensional detector, 141 ... autosampler unit, 142 ... robot arm, 143 ... claw, 144 ... buffer Container, 145 ... Washing container, 146 ... Waste liquid container, 147 ... Sample container, 151 ... Electrophoresis medium filling unit, 152 ... Polymer filling block, 153 ... Syringe, 154 ... Polymer channel, 155 ... Tube, 156 ... Solenoid valve, 161 ... power supply unit, 162 ... high voltage power supply, 163 ... anode buffer container, 64 ... Anode electrode, 501 ... Capillary end face, 502 ... Fluorescent condensing lens, 503 ... Prism, 504 ... Image forming lens, 505 ... CCD, 506 ... Optical axis of fluorescent condensing lens, 507,508 ... in the X-axis direction Two capillaries arranged side by side, 509, 510... Plane formed by the image formation point of light, 511... The optical axis of the image formation lens, 512. Angle θ formed by the CCD surface, 601..., An image obtained by separating light emitted from one capillary, 701... End face of capillary array, 702... Optical axis of fluorescent condensing lens, 703. 705, 803 ... capillary axis, 801 ... capillary array end face, 802 ... optical axis of condensing lens, 805 ... condensing lens, 901 ... end of capillary array, 902 ... CCD light receiving surface, 903 Imaging lens, 904... Optical axis of imaging lens, 905, 906... Spectroscopic fluorescence, 907. Wedge-shaped transparent glass plate, 1000. End of capillary array, 1001... Four band pass filter groups rotating, 1002. Transmission bandpass filter, 1003... Yellow transmission bandpass filter, 1004... Green transmission bandpass filter, 1005... Blue transmission bandpass filter, 1010... Imaging position correction glass plate group, 1020. Laser light, 1405 ... cylindrical convex lens, 1406 ... cylindrical concave lens.

Claims (11)

A plurality of capillaries capable of being filled with an electrophoresis medium;
A power source for electrophoresis of the sample in the electrophoresis medium;
An excitation optical system for irradiating the separated sample with excitation light;
A light receiving optical system including a condensing lens, an imaging lens, a light receiving surface for receiving an image formed by the imaging lens, and a two-dimensional image sensor for detecting fluorescence from a sample emitted from a plurality of capillary ends. When;
An electrophoresis apparatus comprising:
The electrophoretic device , wherein the light receiving surface is inclined with respect to a plane perpendicular to the optical axis, and a plane formed by a plurality of capillary ends is inclined with respect to the optical axis of the condenser lens.   2. The electrophoresis apparatus according to claim 1, wherein the optical axis of the condenser lens is substantially parallel to the capillary axis in the vicinity of the capillary end.   2. The electrophoresis apparatus according to claim 1, wherein a plane formed by a plurality of capillary ends is substantially perpendicular to a capillary axis in the vicinity of the capillary ends.   2. The electrophoresis apparatus according to claim 1, wherein the light receiving optical system includes wavelength dispersion means.   2. The electrophoretic device according to claim 1, wherein the light receiving optical system includes a prism.   2. The electrophoresis apparatus according to claim 1, wherein the sample emits fluorescence having a plurality of wavelengths. A plurality of capillaries capable of being filled with an electrophoresis medium;
A power source for electrophoresis of the sample in the electrophoresis medium;
An excitation optical system that irradiates excitation light to the separated sample;
A light receiving optical system including a condensing lens and a two-dimensional image sensor for detecting fluorescence from a sample emitted from a plurality of capillary ends;
An electrophoresis apparatus comprising:
An electrophoretic device having a wedge-shaped substantially transparent member on a condensing lens side of a two-dimensional image sensor.   8. The electrophoresis apparatus according to claim 7, wherein the wedge-shaped substantially transparent member has a longer wavelength side thickness thinner than a shorter wavelength side thickness.   8. The electrophoresis apparatus according to claim 7, wherein the light receiving optical system includes wavelength dispersion means.   8. The electrophoresis apparatus according to claim 7, wherein the light receiving optical system includes a prism.   8. The electrophoresis apparatus according to claim 7, wherein the sample emits fluorescence of a plurality of wavelengths.

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