JP2640704B2 - Fluorescent pattern reader - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a fluorescent pattern reading apparatus, and more particularly to a fluorescent pattern reading apparatus for detecting a displacement of an optical axis of an optical scanning system for a sample having various thicknesses and a sample having various optical refractive indexes. The present invention relates to a fluorescence pattern reading device that improves fluorescence reading sensitivity by making corrections.

[0002]

2. Description of the Related Art In general, electrophoretic analysis using radioactive isotopes is used to analyze various gene structures including diagnosis of genetic diseases and structural analysis of proteins such as amino acids. In such an electrophoretic analysis method, a fragment of a sample labeled or substituted with a radioisotope is subjected to electrophoresis using a gel, and the distribution pattern of the fragment of the sample developed by electrophoresis is analyzed. Is a method of analyzing a sample by

As an example of reading and analyzing an electrophoresis pattern, a diagnosis of a genetic disease will be described as an example. Human genomic DNA is
Consists of about 3 × 10 ⁹ base pairs. Although its nucleotide sequence is approximately constant throughout the human population, there are variations among individuals when viewed in detail. Such variation in the DNA sequence is called a DNA polymorphism. DN
The polymorphism of A can be found in both the gene region and the non-gene region, but the polymorphism in the gene region often appears as a polymorphism of a protein that is its expression form. Such as blood type, tissue-compatible antigen, and skin color and hair color difference between races.
The various varieties found in the human population derive from this polymorphism. DNA polymorphisms are defined as the germline D
Mutations in NA have accumulated in the population. Such mutations occur at sites with functions important to the presence of humans, and the resulting phenotype, which indicates some pathological condition, is called a "genetic disease". It is said that more than 3,000 genetic diseases exist in the human population.

[0004] The etiology of a genetic disease is an abnormality that occurs on DNA, but by the time it is recognized as a disease, DNA →
There are various stages of mRNA → protein → phenotype (disease). Diagnosis as a disease is usually performed at the last level, but if the above-mentioned flow has a simple linear relationship, diagnosis can be performed at the protein or DNA level.

[0005] The technique underlying DNA diagnosis is Southern.
This is called blotting. This method is basically divided into the following steps. (Step 1) Extraction of sample DNA, (Step 2) Decomposition of DNA by restriction enzyme, (Step 3) Fractionation by molecular weight of DNA using gel electrophoresis, (Step 4) To a thin film filter of fractionated DNA (Step 5) Formation of a hybrid (hybridization) with a previously prepared probe DNA (a DNA having a sequence homologous to the gene to be detected is labeled with an isotope), (Step 6) Autoradiography Detection of hybrids by the above six steps. A nylon membrane or a nitrocellulose membrane is used for the thin film filter in the fourth step.

[0006] In the case of a genetic disease, the organ for extracting the sample DNA is not limited. Usually, leukocytes are separated from a few milliliters of peripheral blood and DNA is extracted therefrom. Steps 1 to 6 usually require about 5 days. In the diagnosis of a genetic disease, based on the above steps, fractionation patterns of normal bodies are obtained and compared. If the patterns are the same, it is determined that the pattern is normal.

[0007] Recently, a probe labeled with a fluorescent dye has been used instead of a radioactive isotope for safety and other problems.
Attempts have been made to excite the fluorescent dye and read an electrophoretic pattern. In the diagnosis of genetic diseases and the determination of the base sequence of DNA, the amount of sample is on the order of 10 ^-15 mol. To obtain a signal-to-noise ratio equivalent to that of a radioactive isotope, advanced optical and signal processing techniques are required. Need.

As an apparatus for detecting a minute sample by fluorescent labeling, there is an electrophoresis apparatus described in JP-A-61-62843. Next, an electrophoresis apparatus using such a fluorescence detection method will be specifically described.

FIG. 15 is an external perspective view of a conventional fluorescent electrophoresis apparatus. Referring to FIG. 15, the electrophoresis apparatus includes: an electrophoresis measurement apparatus 51 that performs electrophoresis of a sample and measures the distribution of fluorescence; a data processing apparatus 52 that performs data processing based on the measured data; Are connected to each other. Electrophoresis measuring device 5
In 1, a door 51 a is opened to inject a gel serving as a base for performing electrophoresis of DNA fragments, and a predetermined amount of a sample to be subjected to electrophoresis is further injected. When the door 51a is closed and the electrophoresis start switch on the operation display panel 51b is pressed, electrophoresis starts. When the electrophoresis is started, the operation state of the electrophoresis measurement device 51 is displayed on a monitor provided on the operation display panel 51b. The data measured by the electrophoresis measurement device 51 is transferred to the data processing device 52, and predetermined data processing programmed in advance is performed. The data processing device 52 includes a computer main body 54, a keyboard 55 for inputting a command from a user, a display device 56 for displaying a processing state and a result, and a printer 57 for recording a result of the data processing. It is configured.

FIG. 16 is a block diagram showing the internal configuration of the electrophoresis measuring device. As shown in FIG. 16, the configuration of the electrophoresis measurement device (51; FIG. 15) includes an electrophoresis device section 63 and a signal processing device section 64.
The two parts are put together to form a migration measuring device. The electrophoresis device section 63 includes an electrophoresis section 5 for performing electrophoresis, a first electrode 2a and a second electrode 2b for applying a voltage to the electrophoresis section 5, an electrophoresis section 5 and the electrodes 2a and 2b.
A support plate 3 for supporting the light, an electrophoresis power supply 4 for applying a voltage to the electrophoresis section 5, a light source 11 for emitting light for exciting a fluorescent substance, and a light source for guiding light from the light source 11. Optical fiber 12 and fluorescent light 1 generated from a fluorescent substance
It is composed of a light collector 14 of an optical system that collects and receives the light 3, an optical filter 15 that selectively passes light of a specific wavelength, and an optical sensor 16 that converts the received light into an electric signal. . The signal processing unit 64 includes an amplifier 17 that receives and amplifies the electric signal from the optical sensor 16 and an analog / digital converter that converts an analog signal of the electric signal into digital data.
A digital conversion circuit 18; and a signal processing unit 19 for performing preprocessing such as averaging processing on the digitally converted data.
And an interface 20 for performing an interface process for sending preprocessed data to an external data processing device, and a control circuit 10 for controlling the entire electrophoretic device and the signal processing system. The digital signal OUT output from the signal processing device 64 is sent to a data processing device (52; FIG. 15), where data processing such as analysis processing is performed.

Next, the operation of the electrophoresis apparatus thus configured will be described. Please refer to FIG. 15 and FIG. The door 51a of the electrophoresis measurement device 51 is opened, a gel is injected into the electrophoresis section 5 inside, and a sample of a DNA fragment labeled with a fluorescent substance is further injected. When a switch on the operation panel 51b is operated to instruct the start of electrophoresis, a voltage from the electrophoresis power supply 4 is supplied to the electrophoresis section 5 by the electrodes 2a and 2b, and electrophoresis is started. A sample labeled with a fluorescent substance by electrophoresis is subjected to electrophoresis in lanes 71, 72, 73, and 74 of each sample as shown in FIG. 19, and is collected for each molecular weight of molecules contained in the sample. , And make a band 66 for each. Lighter molecules have a higher migration speed, so that the migration distance in the same time is longer. The detection of these bands 66 is shown in FIG.
As shown in (A), the light from the light source passes through the optical fiber 12 and irradiates the gel on the optical path 61, so that the fluorescent substance of the label collected in the band 66 in the gel becomes fluorescent 13.
Is generated, and the fluorescence 13 is detected.

The fluorescent light 13 emitted here depends on the extinction coefficient, quantum efficiency, excitation light intensity and the like of the fluorescent substance, but contains only a very small amount of about 10 ^-16 mol per band. Therefore, it becomes very weak light. For example,
Explaining the case where fluorescein isothiocyanate is used as the fluorescent substance, the peak value of the excitation wavelength of the excitation light by fluorescein isothiocyanate is 490 nm, and the peak value of the fluorescence wavelength is 520 nm. The molar fluorescence coefficient is 7 × 10 ⁴ mol ⁻¹ · cm ⁻¹ , and the quantum efficiency is about 0.65.

When a fluorescent substance of 10 ^-16 mol is present in one band, calculation is performed on the assumption that an argon ion laser having a wavelength of 488 nm and an output of 1 mW is used as excitation light. The amount of generated fluorescence is 10
Only ¹⁰ / S fluorescent light photons are generated. Therefore, very weak fluorescence must be detected with high sensitivity.

The front view of the electrophoresis section 5 is shown in FIG.
As shown in FIG. 17 (B), the vertical cross-sectional view is composed of a gel 5a such as polyacrylamide and glass supporting plates 5b and 5c for narrowly supporting the gel 5a from both sides. A sample of, for example, a DNA fragment is injected from above into the gel 5a of the electrophoresis section 5, and the first electrode 2a and the second
By applying a migration voltage to the electrode 2b (FIG. 16) and performing electrophoresis, the light emitted from the light source, for example, a laser beam, passes from the optical fiber 12 through the optical path 61 in the gel 5a.
The fluorescent material on the optical path 61 is irradiated. Thereby, the optical path 6
The fluorescent substance existing on 1 is excited to emit fluorescent light 13. The fluorescent light 13 reaches a light collector 14 of an optical system composed of a combination of lenses, and after being collected, a desired fluorescent wavelength is selected by an optical filter 15 and converted into an electric signal by a one-dimensional optical sensor 16. Is done. In the optical sensor 16, in order to efficiently convert weak light into an electric signal, the light is amplified by a factor of 10 ⁴ to 10 ⁵ using an image intensifier or the like, and the image is electrically amplified by a CCD one-dimensional optical sensor or the like. Convert to a signal. The electric signal converted by the optical sensor 16 is amplified to a signal of a desired level by an amplifier 17, converted from an analog signal to a digital signal by an analog / digital conversion circuit 18, and sent to a signal processing unit 19. In the signal processing unit 19, the signal-to-noise ratio (S / N ratio)
Signal processing such as averaging processing is performed in order to improve. The digital signal data processed in this manner is transmitted to the data processing device 5 by the interface 20.
2 is sent.

FIGS. 18A and 18B are diagrams illustrating an example of a fluorescence intensity pattern signal of a DNA fragment sent from the electrophoresis measurement device 51. FIG. For example, FIG.
(A) As shown in FIG.
Is irradiated on the optical path 61 with laser light, the optical path 6
Since the fluorescent substance of the gel existing on 1 is excited and emits fluorescence, this fluorescence is detected in a predetermined detection position for each lane in the direction of electrophoresis 62 over time. Thus, when the band 66 of each lane passes through the position on the optical path 61, the fluorescence is detected, and the pattern signal of the fluorescence intensity in one lane is shown in FIG.
Is detected as shown in FIG. Therefore, when the band 66 passes through the position on the optical path 61, a peak of the fluorescence intensity is obtained. Therefore, the fluorescence intensity pattern signal shown in FIG. 18B is a fluorescence intensity pattern signal of the band 66 in the electrophoresis direction 62.

In the data processing device 52, a computer main body 54
Receives the data of the fluorescence intensity pattern signal of the DNA fragment transmitted from the electrophoresis measurement device 51, and performs data processing for comparing the molecular weight and determining the DNA base sequence from the data of the fluorescence intensity pattern. The arrangement of bases and the like determined by performing the data processing is symbolized and output, displayed on the screen by the display device 56, or printed out by the printer 57.

In the above example, an example of an apparatus using a fluorescent dye as a sample labeling method has been described. However, as another example, after similarly labeling with a fluorescent dye and performing electrophoresis, a fluorescent pattern based on the electrophoretic pattern is obtained. A reading device is disclosed in Japanese Patent Laid-Open No. 1-16.
No. 7649. The apparatus of this other example is different from the type that reads the distribution of the fluorescent pattern that simultaneously passes through the reading unit while performing the electrophoresis as described above, and is a type that reads the fluorescent pattern of the entire migration unit after the electrophoresis is completed. It has become.

[0018]

By the way, with the type in which the fluorescence pattern of the entire electrophoresis section is read after the electrophoresis is completed as described above, various samples can be read. Samples to be read are roughly divided into, for example, DNA injected into a gel and DNA transferred to a membrane filter.

The gel and the membrane filter have different optical refractive indices. Also, the type and thickness of the gel differ depending on the experimental method. Since there is a difference in optical refractive index and thickness depending on the sample in this way, the optical axis from the irradiation of the excitation light to the emission of the fluorescent light passing through the condenser lens and being focused at the entrance of the light receiving unit is shifted. . Further, the optical axis is also shifted due to the variation in the thickness of the gel glass support or the difference in the optical refractive index. Due to these optical axis shifts, the fluorescence to be detected is focused outside the entrance of the light receiving unit, and the amount of light reaching the optical sensor decreases. Therefore, there is a problem that the detection sensitivity of the fluorescent pattern is reduced.

An object of the present invention is to detect and automatically correct the deviation of the optical axis of an optical scanning system for reading samples of various thicknesses and reading samples of various optical refractive indices.
It is an object of the present invention to provide a fluorescence pattern reading device that improves the fluorescence reading sensitivity.

[0021]

In order to achieve the above object, a fluorescent pattern reader of the present invention emits fluorescence by exciting a fluorescent substance labeling a sample after electrophoresis and development of the sample. In a migration pattern reading device that reads a fluorescent pattern that emits light, a light source that emits irradiation light that excites the fluorescent substance of the migration pattern, and scans the irradiation light from the light source along a predetermined optical axis in the thickness direction of the gel. A light scanning mechanism for irradiating, and a light receiving unit for setting a light receiving surface in a direction different from the optical axis of the irradiation light and separating and receiving the fluorescence of the migration pattern from the scattered light on the reading surface according to the spatial positional relationship of the light receiving path A photoelectric conversion unit that photoelectrically converts an optical signal received by the light receiving unit and outputs an electric signal; and performs an integration operation on the electric signal from the photoelectric conversion unit in correspondence with scanning of irradiation light to amplify and sequentially perform An amplifier for outputting an electric signal of the fluorescence from the dynamic pattern, characterized by comprising an optical axis correcting unit that performs electrical signal to correct the optical axis of the irradiation light by receiving an optical axis detection signal from the amplifier.

[0022]

According to the fluorescent pattern reading apparatus of the present invention,
The light source emits irradiation light for exciting the fluorescent substance in the migration pattern. When the light scanning mechanism scans the irradiation light from the light source along the predetermined optical axis and irradiates the gel in the thickness direction, the light receiving unit sets the light receiving surface in a direction different from the optical axis of the irradiation light, and sets a space for the light receiving path. The fluorescence of the migration pattern is separated from the scattered light on the reading surface and received according to the positional relationship. The photoelectric conversion unit performs photoelectric conversion of an optical signal received by the light receiving unit and outputs an electric signal. Then, the amplifier performs an integration operation on the electric signal from the photoelectric conversion unit in correspondence with the scanning of the irradiation light, amplifies the electric signal, and sequentially outputs a fluorescent electric signal from the migration pattern. Here, an optical axis correction unit is further provided.

The optical axis correction unit receives the electric signal from the amplifier as an optical axis detection signal and corrects the optical axis of the irradiation light. The correction of the optical axis of the irradiation light performed by the optical axis correction unit is performed by detecting the deviation of the optical axis with respect to the read samples having various thicknesses and the read samples having various optical refractive indexes, and detecting the optical axis of the optical scanning system. Is an optical axis correction for correcting For this reason, the optical axis correction unit includes, for example, a folding mirror that changes the optical axis of irradiation light from a light source, an optical axis correction actuator that changes the angle of the folding mirror, an actuator driver that drives and controls the actuator, and an actuator. And a control circuit for supplying a drive control signal to the chain driver. The optical axis correction actuator is, for example, a ceramic actuator, and the control circuit is a feedback control circuit, which receives an electric signal from the amplifier as an optical axis detection signal, and in which the intensity of the optical axis detection signal becomes an extreme value. Then, a drive control signal to the actuator driver is generated.

Thus, in the fluorescence detection of the migration pattern to be read, the optical axis of the folding mirror is set at an angle position where the amount of the fluorescence emitted from the sample surface is maximized, and the fluorescence from the sample can be read with high sensitivity. Can be. Therefore, the difference in the optical distance from the sample surface to the focal lens due to the difference in the thickness and the optical refractive index of each sample can be corrected, and the fluorescent pattern can be read with high sensitivity.

[0025]

DESCRIPTION OF THE PREFERRED EMBODIMENTS One embodiment of the present invention will be specifically described below with reference to the drawings. FIG. 1 is a schematic diagram illustrating an overall configuration of a fluorescent pattern reading device according to an embodiment of the present invention, and FIG. 2 is a diagram illustrating a mounting position of an electrophoresis unit mounted on a measurement unit main body of the fluorescent pattern reading device. is there. As shown in FIG. 1, the fluorescent pattern reading device has an electrophoresis unit 1 and a reading unit 6 which are separated from each other to constitute the entire device. The electrophoresis unit 1 includes a migration unit 5 including a gel serving as a base for performing electrophoresis and a gel support for supporting the gel by sandwiching the gel with a glass plate or the like; A first electrode 2a and a second electrode 2b for applying an electrophoretic voltage to the electrophoresis section 5; a support plate 3 supporting the first electrode 2a and the second electrode 2b and supporting the electrophoresis section 5; And an electrophoresis power supply 4 for supplying a voltage. As described above, the electrophoresis section 5 is composed of a gel such as polyacrylamide as a base on which a sample to be subjected to electrophoresis is developed, and a gel support of a glass plate which supports the gel with the gel sandwiched from both sides (described above). (See FIGS. 17A and 17B)

In the electrophoresis unit 1, the electrophoresis unit 5 is mounted, and the DN that electrophoreses from the upper part of the gel of the electrophoresis unit 5.
A fragmented sample such as an A fragment is supplied, and the first electrode 2a and the second electrode 2b are supplied from the power supply device 4 for electrophoresis.
Is applied with electrophoresis to perform electrophoresis. After the electrophoresis in the electrophoresis unit 1, the electrophoresis unit 5 is detached from the electrophoresis unit 1, and then the reading unit 6
And read the electrophoresis pattern.

The reading unit 6 keeps the electrophoresis unit 5 that has performed electrophoresis in the electrophoresis unit 1 as it is (or in a gel state where only the gel is taken out of the electrophoresis unit 5).
It is mounted on the measuring section main body 7, reads an electrophoresis pattern, and performs data processing. As shown in FIG. 1, the reading unit 6 is configured with a measuring unit main body 7 as a main part, and is configured by adding a data processing device 8, an image printer 9, and the like. The data processing device 8 and the image printer 9 perform data processing, image processing, and image processing on the electrophoresis pattern image data read by the measurement unit main body 7.
A determination process is performed, and the read electrophoresis pattern data is processed and output. As shown in FIG. 2, the measuring unit main body 7 includes a reading table 7 c to which an electrophoresis unit (electrophoresis unit unit including a gel and a gel support) 5 that has already electrophoresed in the electrophoresis unit 1 is mounted and read. It is provided directly below the lid 7a at the top of the main body.

The electrophoresis section 5 removed from the electrophoresis unit 1 is attached to the lid 7a of the upper section of the measurement section main body in the measurement section main body 7.
Is opened and attached to the reading table 7c. Reading stand 7c
After the gel to be read by the electrophoresis unit 5 is attached to the
Is closed and the reading start switch on the operation display panel 7b of the measuring section main body 7 is pressed, the measuring section main body 7
Start reading the electrophoresis pattern of the gel. When the reading of the electrophoresis pattern is started, the measurement unit body 7
The scanning of the irradiation light from the point light source built in is started, the excitation light is irradiated on the gel of the mounted electrophoresis section 5 to excite the fluorescent substance, and the emitted fluorescent light is received, and the distribution of the fluorescent substance distribution is Measure the pattern. The data processing device 8 performs data processing based on the read data measured by the measuring section main body 7 and controls the measuring section main body 7. The processed data is output and visualized by the image printer 9. In this image printer, a migration pattern is printed on each sample while distinguishing the migration pattern.

When performing electrophoretic analysis of a sample using a fluorescent pattern reader, as described above, first, the sample (DNA fragment) labeled with a fluorescent dye (fluorescent substance) is used by using the electrophoresis unit 1. Perform electrophoresis. After completion of the electrophoresis for a predetermined time of about 5 hours, the electrophoresis section 5 is detached from the electrophoresis unit 1. With the gel of the electrophoresis unit 5 removed, with the electrophoresis unit 5 as it is or with the glass of the gel support removed, open the upper lid 7a of the measurement unit main body 7 of the reading unit 6, and open the internal reading table. 7c. Then, close the lid 7a,
The setting to the reading unit is completed. At this time, if the gel on which the electrophoresis has been performed is a sample that is not labeled with a fluorescent dye, a process of applying a fluorescent dye to the sample may be performed at this stage. At this time, processing such as drying of the gel is also performed.

Next, an operation for instructing the start of reading the electrophoresis pattern is performed. The reading start operation is performed according to a start instruction of a pressing operation of a reading start switch of the operation display panel 7b or a reading start instruction from the data processing device 8. When the reading operation is started by the data processing device 8, the mounting state of the electrophoresis unit in the measuring unit main body 7 is sent to the data processing device 8 through a control signal line, and the state is confirmed.
Controls the operation of the reading unit of the measuring unit body 7. In this case, the parameter setting such as the reading speed at the time of operation is registered in the data processing device 8 in advance, so that the reading start operation is performed automatically, so that the burden on the switch operation of the operator is reduced. Is done.

The read distribution data of the fluorescent dye is sent to the data processing device 8. The data processing device 8 performs desired processing that is programmed in advance, such as processing for detecting the peak of the fluorescence intensity and processing for obtaining the migration distance. The data resulting from the data processing is printed out by the image printer 9 as a necessity, in the form of a gray-scale image of the fluorescence intensity, or printed out as an image in which the fluorescence intensity is divided into contour lines or colors or densities. An image printed and output as a gray-scale image corresponding to the fluorescence intensity is an image similar to a radioactive X-ray film image obtained by labeling with a conventionally used radioactive substance and performing electrophoresis. If necessary, the data resulting from the data processing is stored as digital data in a magnetic or optical recording device.

FIG. 3 is a block diagram showing the configuration of the main part of the measuring section main body including the optical axis correcting mechanism section. FIG. 4 is a diagram illustrating a schematic configuration of a mechanism of an optical axis correction mechanism that performs automatic optical axis correction, and FIG. 5 is a diagram illustrating a configuration of an actuator of the mechanism of the optical axis correction mechanism. As shown in the upper part of the block diagram of FIG.
An optical axis correction mechanism 40 that performs optical axis correction of irradiation light is provided. The schematic configuration diagram of the mechanical part of the optical axis correction mechanism shown in FIG. 4 is a longitudinal sectional view of a part of the structure of the optical system of the mechanical part of the optical axis correction mechanism 40 shown in FIG.

First, with reference to FIG. 3, the flow of the whole operation in the measuring section main body 7 for reading the fluorescent pattern will be described. In the measurement unit main body, a laser beam 31 for exciting a fluorescent substance that labels a migration pattern after electrophoresis is emitted from a light source 21 and a mirror driver 30 is emitted.
The light is irradiated onto the turning mirror 44 of the optical axis correction mechanism 40 while being scanned in the right and left directions in FIG. During the operation of correcting the optical axis of the irradiation light (laser beam), the folding mirror 44 is driven by the actuator 41 attached to the folding mirror 44 to displace the folding mirror 44 so that the laser beam 3
The optical axis 1 is changed in the front and back direction of the drawing, and the displacement of the return mirror 44 is positioned at a position where the sensitivity of receiving the fluorescence of the electrophoresis pattern from the gel 5 becomes optimal. Thereby, the laser beam 31 scanned by the vibrating mirror 22
Is reflected by the folding mirror 44 displaced for optical axis correction, and then irradiated in the thickness direction of the gel of the electrophoresis unit 5.

That is, the laser beam 31 from the light source 21
Is reflected by the oscillating mirror 22 so as to be scanned in the horizontal direction of the drawing, further reflected by the folding mirror 44, and irradiates the gel of the migration unit 5 to be read. Each time the laser beam 31 is scanned once, the entirety of the electrophoresis unit 5 is moved by moving the laser beam relative to the electrophoresis unit 5 in a direction perpendicular to the scanning direction of the laser beam 31, that is, in the front and back directions in the drawing. Can be read. The movement in the sub-scan direction is performed by the reading table 7.
This is carried out by moving the electrophoresis section 5 placed on c by a reading drive mechanism (FIG. 2). Further, an optical trap 32 is provided on the opposite side of the irradiation surface of the electrophoresis unit 5 from which the laser beam 31 is irradiated, so that the laser beam 31 transmitted through the gel does not adversely affect the stray light.

The irradiation of the spot light of the laser beam 31 scanned by the vibrating mirror 22 causes the fluorescence 13 emitted from the gel of the electrophoresis section 5 to be received through the condenser 23. The condenser 23 receives the fluorescent light 13 so that the optical axis of the light receiving path is different from the optical axis of the spot light illuminating the electrophoresis unit 5 and the spatial positional relationship of the optical path of the light receiving path is determined by an optical lens system. , An optical path is configured, the detection sensitivity from the scattered light emitted from the irradiation surface of the electrophoresis unit 5 is increased, and the fluorescence 13 is received. The fluorescent light 13 received and collected by the light collector 23 is further photoelectrically converted by a photoelectric conversion unit 24 including a condenser lens, an optical filter, and a photomultiplier, and is converted into an electric signal. The converted electric signal is input to the amplifier 25. Then, the electric signal amplified by the amplifier 25 is input to an analog / digital converter (hereinafter abbreviated as A / D converter) 26 and converted into digital data. The detection signal converted into digital data is stored in the memory 28, and the data stored in the memory 28 is sent to the data processing device 8 through the interface 29. The overall control of such a series of signal processing is performed by the control circuit 27.

Next, the operation of the optical axis correcting mechanism 40 will be further described with reference to FIG. FIG. 4 shows a schematic configuration of the optical axis correction mechanism 40 centering on the optical system. In the fluorescence pattern reading device here, the gel 5 of the electrophoresis section 5 is used.
Since a is read for each glass plate (such as the glass support plate 5b and the glass plate of the reading table 7c), the difference in the thickness of the glass and the unevenness in the gel thickness are also considered.
The optical distance differs for each sample depending on the thickness of the gel used, the difference in the refractive index, and the like. For this reason, the focus position of the fluorescent light 13 received by the light collector 23 shifts, and the amount of received light differs for each sample. For this reason, by the operation of the optical axis correcting mechanism 40 here, the folding mirror 44 is displaced by the actuator 41 and the gel 5 of the electrophoresis section 5 is displaced.
As shown in FIG. 4, the irradiation position of the laser beam irradiated on the laser beam a is shifted in the left-right direction, so that the optical axis 45 from the return mirror 44 to the condenser 23 is corrected to an optimum position, and the gel 5 a Automatic optical axis correction is performed so as to stably receive the fluorescent light 13 from the camera.

For optical axis correction, ordinary optical techniques in an optical system can be used. For example, a correction method using an acousto-optic element using a light diffraction phenomenon or a method using another actuator is used. In this embodiment, a method in which the folding mirror 44 is controlled by the actuator 41 using a piezoelectric ceramic element is used.

Referring again to FIG.
1 will be described. The fluorescent light 13 generated from the gel is received and collected by a light collector 23, converted into an electric signal by a photoelectric converter 24, amplified by an amplifier 25, and further converted into digital data by an A / D converter 26. Therefore, this digital data is used as a detection signal for generating a control signal for optical axis correction. That is, digital data proportional to the detected fluorescence intensity is used as an optical axis detection signal, and a feedback loop control system is configured so that the detected signal intensity is maximized. In the processing of this control system, a macro computer constituting the control circuit 27 performs a series of control signal processing and sends a control signal to the actuator control circuit 43, whereby a control operation is performed. The actuator control circuit 43 sends a control signal to the actuator driver 42, and the actuator driver 42 drives the actuator 41. A folding mirror 44 is attached to the actuator 41.
When 1 is driven by the actuator driver 42, the reflection angle of the return mirror 44 is displaced. The optical axis of the laser beam applied to the electrophoresis unit 5 changes due to the minute displacement of the folding mirror 44. The change in the optical axis is controlled so that the magnitude of the optical axis detection signal is maximized.

In the control operation of the actuator for displacing the folding mirror 44, first, when the gel of the electrophoresis section 5 is read, a laser beam is scanned, and the voltage applied to the actuator 41 is changed every time scanning is performed. The mirror 44 is displaced. The data of the magnitude of the electric signal detected from the photoelectric conversion unit 24 every time scanning is performed is received via the amplifier 25, the A / D converter 26, and the memory 28, and is sequentially compared. The voltage applied to the actuator 40 when? A control signal for generating a voltage applied to the actuator when the magnitude of the electric signal is maximum is used as a reference control signal for setting the optimum position of the folding mirror 44. When changing the voltage applied to the actuator, first, the variable width and the step width are set. First, the variable width and the step width of the applied voltage are set to be large, and the value of the applied voltage at which the electric signal is high is roughly grasped. Then, the value of the applied voltage obtained next is set as a reference, the variable width and the step width are set small, and the applied voltage is determined. By supplying the applied voltage determined in this manner to the actuator 41, the control of the optical axis correction of the laser beam 31 as the irradiation light is performed.

Next, the actuator 41 of the optical axis correcting mechanism 40 will be described. Referring to FIG. 5, there is shown a structure of an actuator 41 to which a folding mirror 44 of a mechanism of the optical axis correcting mechanism is attached. As the actuator 41 constituting the main part, as shown in FIG. 5, a piezoelectric actuator having a structure in which two piezoelectric ceramic plates 41a and 41b made of a piezoelectric material are bonded to each other is used. The piezoelectric type actuator 41 here is called a bimorph type actuator. The bimorph type actuator is characterized by a large displacement enlargement ratio in addition to a simple manufacturing method of bonding two ceramic plates with an adhesive. is there. Basically, it has a structure in which two piezoelectric ceramic plates that expand and contract in the length direction are bonded together, and when one expands, the other contracts, causing a bending displacement as a whole. To increase the response frequency and generate the driving force,
An elastic plate 41c called a shim is sandwiched between the ceramic plates 41a and 41b. As the elastic plate 41c, a phosphor bronze plate or the like is used. In the bending operation, an electric field is applied using the shim (elastic plate) 41c as a negative pole and the two ceramic plates 41a and 41b as positive poles. The bending mirror 4 to which the actuator 41 is attached is formed by the applied bending displacement of the actuator 40.
4 are displaced.

As described above, the mechanical portion of the optical axis correcting mechanism 40 has the end of the shim 41c sandwiched between the two ceramic plates 41a and 41b of the actuator 41 attached to the back side of the mirror surface of the folding mirror 44, and is folded back. The end face of the actuator 41 on which the mirror 44 is not attached is fixed. The shim 41c here is bent in a U-shape. This allows the mirror 4 to be folded without bending the shim.
4, a larger displacement can be obtained. Regarding the drive control of the actuator 41,
The variable speed of the applied voltage and the scan speed of the laser beam 31 are synchronized.

Next, an optical scanning mechanism for irradiating the laser irradiation surface of the electrophoresis section 5 with the laser beam will be described. FIG. 6 is a diagram illustrating an optical scanning mechanism that scans the gel surface with a laser beam using a vibrating mirror, and FIG. 7 is a diagram illustrating the relationship between the rotation angle of the vibrating mirror and the moving distance of the spot light of the laser beam. It is. In the optical scanning mechanism here, the electrophoresis section 5
And the arrangement positions of the light source 21 and the vibrating mirror 22 are in a positional relationship as shown in FIG.
When the mirror 2 is driven by the mirror driver 30 so as to vibrate at a constant angular velocity, the moving speed of the light spot at both ends of the electrophoresis unit 5 is faster than that near the center (X = 0). Therefore, the detection sensitivity of the fluorescence detected from the sample of the electrophoresis unit 5 is different between the center and the end. On the other hand, in the present embodiment, the moving speed of the laser spot light on the gel of the
The speed of driving the oscillating mirror is corrected and controlled. That is,
The relationship of the mirror angle θ to the spot light position X is
The relationship is as shown in FIG. 7. When the distance Z between the center of rotation of the vibrating mirror and the center of the electrophoresis unit 5 is used, the mirror angle θ is expressed by the following equation. θ = arctan (X / Z) where Z is the distance from the rotation center of the vibrating mirror 22 to the gel of the migrating unit 5, and X is the perpendicular from the rotation center of the vibrating mirror 22 to the gel surface of the migrating unit 5. This is the distance in the plane direction of the gel with the lowered point as the origin.

As a method for correcting the relationship between the rotation angle and the moving distance in this type of optical scanning mechanism, fθ
Although there is a method using a lens, the fθ lens is expensive and the apparatus becomes heavy because the fθ lens is mounted. Therefore, here, the correction between the rotation angle of the vibration mirror and the moving distance of the optical scanning mechanism is performed by a mirror driver. 30 is provided with a control circuit for variably controlling the rotational angular velocity of the vibrating mirror 22;
This is performed by correcting and controlling the rotational drive speed of No. 2.

FIG. 8 is a block diagram showing a main configuration of a control circuit of a mirror driver for controlling the rotational driving of the oscillating mirror. The galvanometer scanner is used as the actuator of the oscillating mirror.
Control is performed by applying a voltage proportional to the rotation angle. In order for the spot light of the laser beam to move at a constant speed on the irradiation surface of the gel, the distance X of the irradiation surface and the time t may be controlled so as to be in a proportional relationship. Since the relationship between the rotation angle θ of the vibrating mirror and the moving distance X of the spot light is as shown in FIG. 7, the horizontal axis of the graph in FIG. 7 corresponds to the time axis, and the vertical axis corresponds to the voltage axis. A signal having the generated voltage waveform is generated, and this is used as a drive control signal for driving the vibrating mirror. The generation of such a drive control signal is performed by a control circuit in the mirror driver 30, and the generated drive control signal is supplied to the actuator of the vibration mirror 22 to control the drive of the vibration mirror 22.

As shown in FIG. 7, the mirror driver 30 includes a read-only memory 30a storing function waveforms,
A digital converter that converts the read function data into a voltage signal
An analog conversion circuit 30b, a driver 30c for amplifying the converted voltage signal and outputting it as a drive control signal, a counter 30d for providing a read address to the memory in time series, and an oscillation circuit 30e for providing a clock signal to the counter. It is composed of

The oscillation circuit 30e operates according to an instruction from the control circuit 27 of the measuring section main body, and a clock signal from the oscillation circuit 30e is input to the counter 30d.
d counts the clock signal, and the read only memory 3
A read address to be supplied to 0a is generated in time series. When the read addresses sequentially generated from the counter 30d are supplied in time series to the read-only memory 30a, function data stored in advance is sequentially read from the read-only memory 30a. Function data (FIG. 7) relating to the rotation angle of the oscillating mirror is written in advance in the read-only memory 30a, and such function data is read out in a time-series manner. In this example, the number of bits of the function data is 12 bits. The read function data is converted into a voltage signal of an analog signal for controlling the rotation angle of the oscillating mirror in the digital / analog conversion circuit 30b. This voltage signal removes step-like noise by filtering in the driver 30c, further amplifies the power, and provides a drive control signal as a drive control signal.
Supplied to Thus, the vibrating mirror can be vibrated at a desired rotation angular velocity such that the moving speed (scan speed) of the spot light of the laser beam in the electrophoresis unit is constant.

The scan speed here is set to be 0.5 Hz, 1 Hz, 2 Hz, 5 Hz, 10 Hz, 20 Hz so as to be approximately equally logarithmic.
The configuration is such that it can be varied at each speed of Hz, 50 Hz, 100 Hz, and 200 Hz. This is because the reading speed can be changed in accordance with the amount of the fluorescent substance labeled on the sample to be electrophoresed or the difference in the quantum efficiency of the fluorescent substance, so that the reading can be performed efficiently. In this case, the scanning speed is specified from the operation display panel 7b or the data processing device 8. From the control circuit 27 to the mirror driver 30,
When the scan speed instruction data is sent, the counter 30
By controlling d and the oscillation circuit 30e, the oscillating mirror 22 is driven at a desired scanning speed.

As described above, the laser beam from the light source 21 is scanned by the driving control of the vibrating mirror 22, and is radiated in the electrophoresis unit 5 as spot light moving at a constant speed. As a result, the fluorescent substance of the gel of the electrophoresis section 5 in the portion irradiated by the irradiation light of the laser light is excited, and emits fluorescence 13 (FIG. 3).

FIG. 9 is a view showing the configuration of the main part of the light collector for receiving the fluorescence generated from the gel and the photoelectric conversion unit, focusing on the optical path. As described above, the gel 5a of the electrophoresis section
Are supported between glass gel supports 5b and 5c. Here, as the gel supports 5b and 5c, in the case of the electrophoresis section 5 of this example, borosilicate glass having relatively little fluorescence is used for the gel supports 5b and 5c. In addition,
As the gel supports 5b and 5c, quartz glass or various optical glasses can be used.

In the electrophoresis section 5, when the irradiation light of the scanned and moving laser beam 31 is applied, the irradiation light of the laser beam 31 passes through the gel supports 5c, 5a and 5b in the thickness direction. , Reaches the gel 5a.
In the gel 5a, the light of the laser beam 31 travels in the thickness direction. The thicknesses of the gel support plates 5b, 5c and the gel 5a are about 5 mm and about 0.35 mm, respectively, and the light of the laser beam 31 applied in the thickness direction of the gel support plates 5b, 5c and the gel 5a The intensity of the light reaching the gel at any position of the part 5 is substantially equal. In addition, the spread and decrease in intensity of the laser beam 31 due to light scattering generated on the light incident surfaces of the gel 5a and the gel supports 5b and 5c are greatly reduced because the excitation light is incident perpendicularly to the surface in the thickness direction. Less. The laser beam 31 that has passed through the gel 5a enters the optical trap 32 and is attenuated so as not to adversely affect as stray light.

As described above, the scanning of the laser beam 31 irradiates the irradiation light onto the gel 5a, and the fluorescence generated from inside the gel 5a by the irradiation of the irradiation light (fluorescence excitation light) is converted into the excitation light. The light is collected by the light collector 23 together with the scattered light by itself. The scattered light generated in the gel supports 5b and 5c is geometrically and optically separated by forming an optical path as shown in FIG. 7 and spatially positional relationship of the light receiving path, and only the fluorescence from the gel is extracted. And sent to the photoelectric conversion unit 24. In the photoelectric conversion unit 24,
The scattered light and the fluorescence generated in the gel are separated using an optical filter, and the weak fluorescence is converted into an electric signal by the photomultiplier. The configuration of the optical path of the condenser 23 is as follows.
As shown in FIG. 9, the light is introduced into the photoelectric converter 24 by the optical fiber array 23b in which the light entrances are integrated into one.

The configuration of the optical system in the condenser 23 and the photoelectric conversion unit 24 will be described with reference to FIG.
As shown in FIG. 9, an optical path is configured so that the fluorescence from the gel 5a of the electrophoresis section 5 and the scattered light of the excitation light generated from the gel supports 5b and 5c are received by the cylindrical lens 23a and collected. I have.

Referring to FIG. 9, the fluorescence 1
The scattered light of the excitation light generated from 3 and the gel supports 5b and 5c reaches the cylindrical lens 23a, and the scattered light and the fluorescent light are imaged on the opposite side by the cylindrical lens 23a as shown in the figure. A in the figure
The point is the focus for the fluorescence from the gel 5a and the scattered light of the excitation light generated from the gel 5a. In the case of the scattered light of the excitation light generated on the surfaces of the light incident surfaces of the gel supports 5b and 5c, for example, in the case of the scattered light from the surface of the gel support 5c, an image is formed at point A 'in the figure. . Here, the optical fiber array 23b is disposed at the position of the image forming point A so that only the fluorescence from the gel 5a is received at the entrance of the light, so that the optical fiber array 23b has a geometrical optical relationship due to the spatial positional relationship of the light receiving path. To
The fluorescence is separated from the scattered light from the gel support.

As described above, the method of irradiating the excitation light in the thickness direction is such that the refractive index between the gel and the glass as the gel support is relatively close to about 1.4 to 1.5, and the gel and the gel support are used. Due to the close contact of the body glass interface, very little scattered light is generated at this interface. Therefore, the light received at the point A has less scattered light of the excitation light generated on the surface of the gel 5a,
Only the fluorescence from the inside of the gel 5a is largely received.

When one of the gel supports 5b and 5c or both of the gel supports is removed and the irradiation light as excitation light is directly scanned on the gel 5a, the gel as described above is used. There is a possibility that almost the same amount of scattered light as the amount generated from the glass surface as the support is generated from the gel surface. In this case, the reading table (7c; FIG. 2) of the measurement unit main body is placed. Since the thickness of the glass has the same effect as the thickness of the glass of the gel support as described above, the fluorescence from the gel surface can be reliably detected without lowering the detection sensitivity. The gel supports 5b, 5
When one or both of c is removed, at this point, a treatment for coloring the gel 5a with a dye or the like is performed. In particular, when it is not necessary to remove the gel supports 5b and 5c, reading the gel 5a with the gel supports 5b and 5c attached can improve the signal-to-noise ratio.

In the configuration of the condenser 23, only one cylindrical lens 23a is used. However, the position is symmetrical with respect to the scanning plane of the laser beam, and further, the opposite side of the sample. A cylindrical lens may be mounted on the camera. In particular, when the amount of fluorescence to be detected is insufficient, for example, a cylindrical lens and an optical fiber array are placed in four directions surrounding a scanning line of the gel that generates fluorescence, and the fluorescence that emits weakly is set. Is condensed to increase the amount of detected light. In that case, it is effective to shift the optical axis so that the surface reflections of the opposed cylindrical lenses are not affected by each other.

The fluorescent light collected by the entrance (point A) of the optical fiber array is guided into the optical fibers of the fiber array 23 and supplied to the photoelectric conversion unit 24 from the light exits where the optical fibers are bundled. Is done. The fluorescence input to the photoelectric conversion unit 24 is transmitted to the first lens 24a, the aperture 24b, and the second lens 24a.
Only the parallel component is extracted using the lens 24c, and is incident on the optical filter 24d. And the optical filter 2
4d, the scattered light component is removed, and the third lens 24
The light is condensed by e, guided to the photomultiplier tube 24f, and the detected fluorescence is converted into an electric signal.

As described above, in the photoelectric conversion unit 24, only the parallel light component is incident on the optical filter 24d by the second lens in order to improve the wavelength separation of the optical filter 24d, and the light is incident on the optical filter 24d at right angles. Then, the scattered light of the excitation light generated in the gel is separated by the optical filter 24d to improve the signal-to-noise ratio.
And the light is guided to the photomultiplier tube 24f.

After the fluorescent light is received and condensed as described above, and the scattered light is removed by the optical filter, the fluorescent light guided to the photomultiplier tube 24f is converted into an electric signal by the photomultiplier tube 24f. The converted electric signal is input to the amplifier 25. Then, in the amplifier 25, the weak signal is sufficiently amplified by the amplification stage including the integration circuit.

FIG. 10 is a circuit diagram showing a configuration of an amplifier including an integrating circuit. As shown in FIG. 10, the amplifier 25 is provided with an integrating circuit composed of an operational amplifier 25a at the preceding stage and an output amplifier circuit composed of an operational amplifier 25b at the next stage. Make up. The electric signal from the photomultiplier tube 24f is supplied to an operational amplifier 25a.
Is input to The operational amplifier 25a constitutes an integrating circuit together with the capacitor 25c and the switch 25d for controlling the integrating operation. The output of the integrating circuit is input to the following operational amplifier 25b and is amplified by a gain determined by an external resistor. It is sent to the next analog-to-digital conversion circuit.

The operation of the amplifier 25 including the integrating circuit configured as described above will be described with reference to the timing chart of FIG. Since the output of the photomultiplier tube 24f has a very large output impedance, it can be almost regarded as a current source. The operational amplifier 25a has an FET
(Field effect transistor) An input type high input impedance is used, and when the switch 25d is turned off, the output current ip of the photomultiplier tube 24f is output.
Becomes the current flowing through the capacitor 25c as it is. With this current, the output voltage of the operational amplifier 25a becomes a ramp function output as shown in FIG. In this integration operation, integration is performed for a time corresponding to one pixel, and a sampling circuit in the analog-to-digital conversion circuit 26 performs sampling in accordance with the timing of the S / H clock, and holds it as it is. The conversion circuit 26 converts the signal into a digital signal.

After being held, the charge stored in the capacitor 25c is discharged by activating the C / D clock which is the C / D control signal applied to the switch 25d. Hereinafter, similarly, the integration operation for such a time corresponding to one pixel is repeated. An amplification stage using an integration circuit using such an operational amplifier differs from a pseudo integration circuit consisting only of a resistor and a capacitor, in that the charge from the photomultiplier tube 24f can be almost completely integrated. Therefore, a high signal-to-noise ratio can be obtained. Also, the integration time can be arbitrarily changed by changing the C / D clock of the C / D control signal for the switch 25d. Therefore, it is possible to easily adjust the degree of amplification for amplifying the weak signal comprehensively. In the case of this example, by synchronizing with the scanning operation of the mirror driver 30 shown in FIG. 6, it is possible to control according to the size of the area of the sample to be read, and it is possible to eliminate unnecessary reading time. it can. Further, since the scan speed of the irradiation light (excitation light) and the integration time of the amplifier on the light receiving side can be freely set in accordance with the intensity of the fluorescence from the sample, a very flexible apparatus can be configured.

As described above, the electric signal amplified by the amplifier 25 (FIG. 3) is transmitted to the next analog / digital conversion circuit 26.
And converted into digital data. The fluorescence detection signal converted into digital data is stored in the memory 28, and the data stored in the memory 28 is sent to the data processing device 8 through the interface circuit 29. The control circuit 27 performs overall control of such a series of signal processing.

Next, a description will be given of a modification of the components in the fluorescent pattern reading apparatus of the present embodiment. Another example of the actuator 41 in the optical axis correction mechanism 40 will be described. FIG. 12 shows another configuration example of the actuator in the optical axis correction mechanism. As shown in FIG. 12, an actuator 46 according to another configuration example has an elastic shim 4 sandwiched between two ceramic plates 46a and 46b.
6c is an actuator having a flat structure. The mechanism of the optical axis correction mechanism 40 includes a folding mirror 44.
The actuator 46 is attached in the side direction from the back side of the actuator. The inverse piezoelectric effect of the voltage applied to the actuator 46 causes a distortion proportional to the applied voltage, and the effect of the distortion is used as a driving force. Use
The folding mirror 44 is displaced. As with the actuator 41 (FIG. 5) described in the previous embodiment, the shim 41c
A large displacement can be obtained in the type in which the displacement is bent into a U-shape, but it may be difficult to accurately control the displacement in proportion to the applied voltage. On the contrary,
In the piezoelectric actuator 46 according to the other configuration example, the shim 4
The folding mirror 44 is attached without bending 6c. Accordingly, in the optical axis correction mechanism using the piezoelectric actuator 46, it is possible to accurately control the displacement amount in proportion to the applied voltage. When a piezoelectric actuator is used as an actuator for obtaining a driving force for displacing the folding mirror 44, in order to improve the response,
When the driving force is insufficient, a configuration may be employed in which two piezoelectric actuators are used to obtain a driving force for displacing the folding mirror 44.

FIG. 13 is a diagram showing another example of the structure of the optical axis correcting mechanism constituted by using two piezoelectric actuators. The two actuators according to this configuration example are actuators having a structure in which the shim of an elastic plate sandwiched between two ceramic plates is flat like the actuator 46 shown in FIG. The optical axis correcting mechanism includes a plurality of actuators 4
In the mechanical section having the structure in which the mirror 6 is attached,
Each actuator 46 must be driven at the same displacement and the same frequency so as to balance the left and right of the displacement of the actuator, but a large driving force can be obtained with a relatively small actuator.

Further, in still another configuration example shown in FIG.
The mechanism of the optical axis correction mechanism is constituted by using the four piezoelectric actuators. The four actuators according to this configuration example are configured using an actuator 47 having a structure in which a shim of an elastic plate sandwiched between two ceramic plates is bent in a U-shape. In such a mechanism unit in which four actuators 47 are attached to the folding mirror 44, it is necessary to drive each actuator 47 so that the displacement of the folding mirror 44 can be balanced in the front-rear and left-right directions. A large driving force can be obtained with a relatively small actuator, and the displacement of the folding mirror 44 can be finely adjusted by individually adjusting the voltage applied to each actuator 47.

Next, another modification of the optical axis correcting mechanism will be described. In the overall optical axis correction control system of the optical axis correction mechanism described with reference to FIG.
Feedback control for correcting the deviation of the optical axis from the optimal detection position is performed. In this case, for example, an optical axis correction mechanism that corrects the optical axis by moving the side of the light collector 23 is used. It may be. That is, the electrophoresis section 5
If the optical axis of the laser beam applied to the mirror cannot be corrected due to the displacement of the mirror 44 by the actuator 41, the focus position of the fluorescent light received by the collector 23 deviates from the light receiving surface position B of the collector 23. . For this reason, here, the optical axis 45 is corrected by providing a drive mechanism for directly moving the light collector 23 side. For example, by attaching an actuator to the condenser 23, the condenser 23 is displaced in a direction to detect more fluorescence to be received. Further, the direction and arrangement position of the cylindrical lens 23a in the condenser 23 may be changed. Further, the light receiving surface position B of the light collector 23 may be moved. In either case, the feedback control is performed to displace the direction and the position where more fluorescence to be received is detected. In addition, such a light collector 2
Regarding the actuator control method for displacing the mirror 3, the same control method as that for displacing the folding mirror as described above is used. When an acousto-optic device is used, it goes without saying that a similar effect can be obtained by inserting the acousto-optic device between the galvanometer scanner and the light source and changing the angle of incidence of the laser beam on the galvanometer scanner mirror.

[0070]

As described above, according to the fluorescent electrophoresis pattern apparatus of the present invention, the optical axis position at which the fluorescence detection sensitivity is optimized is automatically set, and the fluorescence of the migration pattern is read. Therefore, various samples can be read with high sensitivity without regard to the thickness and optical refractive index of the sample.

[Brief description of the drawings]

FIG. 1 is a schematic configuration diagram illustrating the overall configuration of a fluorescent electrophoresis pattern reader according to one embodiment of the present invention;

FIG. 2 is a diagram illustrating a mounting position of an electrophoresis unit mounted on a measurement unit main body;

FIG. 3 is a block diagram showing a configuration of a main part of a measurement unit main body including an optical axis correction mechanism;

FIG. 4 is a diagram illustrating a schematic configuration of a mechanism of an optical axis correction mechanism that performs automatic optical axis correction;

FIG. 5 is a diagram illustrating a configuration of an actuator of a mechanism of the optical axis correction mechanism.

FIG. 6 is a diagram illustrating an optical scanning mechanism that scans a gel surface with a laser beam using a vibrating mirror;

FIG. 7 is a view for explaining a relationship between a rotation angle of a vibrating mirror and a moving distance of a spot light of a laser beam;

FIG. 8 is a block diagram showing a configuration of a main part of a control circuit of a mirror driver that rotationally controls a vibration mirror;

FIG. 9 is a diagram showing a detailed configuration of an optical system of a condenser and a photoelectric conversion unit;

FIG. 10 is a circuit diagram showing a circuit configuration of an amplifier including an integrating circuit;

FIG. 11 is a time chart showing the timing of the read operation of the amplifier;

FIG. 12 is a diagram showing another configuration example of the actuator in the optical axis correction mechanism.

FIG. 13 is a diagram showing another example of the configuration of the mechanism of the optical axis correction mechanism configured by using two piezoelectric actuators;

FIG. 14 is a diagram showing another example of the structure of the mechanism of the optical axis correction mechanism configured using four piezoelectric actuators;

FIG. 15 is a perspective view showing the appearance of a conventional fluorescent electrophoresis apparatus,

FIG. 16 is a block diagram showing an internal configuration of the electrophoresis measurement device;

FIGS. 17A and 17B are a front view and a longitudinal sectional view of an electrophoresis section showing an operation principle of electrophoresis pattern detection by a fluorescence method, respectively.

18 (A) and 18 (B) are diagrams illustrating an example of a fluorescence intensity pattern signal of a DNA fragment sent from the electrophoresis measurement device,

FIG. 19 is a diagram showing an example of the distribution of DNA fragments subjected to electrophoresis.

[Explanation of symbols]

 DESCRIPTION OF SYMBOLS 1 ... Electrophoresis unit 2a ... 1st electrode 2b ... 2nd electrode 3 ... Support plate 4 ... Power supply device for electrophoresis 5 ... Electrophoresis unit (electrophoresis unit) 6 ... Reading unit 7 ... Measurement unit main body 8 ... Data processor 9 ... Image Printer 10 ... Control Circuit 11 ... Light Source 12 ... Optical Fiber 13 ... Fluorescence 14 ... Condenser 15 ... Optical Filter 16 ... Optical Sensor 17 ... Amplifier 18 ... Analog / Digital Conversion Circuit 19 ... Signal Processing Unit 20 ... Interface 21 ... Light source 22 Vibrating mirror 23 Condenser 24 Photoelectric conversion unit 25 Amplifier 26 Analog-to-digital conversion circuit 27 Control circuit 28 Storage circuit 29 Interface 30 Mirror driver 31 Laser beam 32 Optical trap 40 Optical axis correction mechanism 41 41 Actuator 42 Actuator driver 43 Actuator Data control circuit 44 ... Folding mirror 46 ... Actuator 47 ... Actuator 51 ... Migration measurement device 51a ... Door 52 ... Data processing device 53 ... Cable 54 ... Computer body 55 ... Keyboard 56 ... Display 57 ... Printer 63 ... Electrophoresis unit 64 ... Signal processing device 61: optical path 62: scanning line 66: band 71, 72, 73, 74: lane.

 ──────────────────────────────────────────────────続 き Continuing on the front page (72) Inventor Jin Fujimiya 6-81 Onoecho, Naka-ku, Yokohama-shi, Kanagawa Prefecture Within Hitachi Software Engineering Co., Ltd. No. 81, Hitachi Software Engineering Co., Ltd. (56) References JP-A-4-36652 (JP, A) JP-A-2-203255 (JP, A) JP-A-61-3043 (JP, A) JP-A-56-122945 (JP, A) JP-A-3-10147 (JP, A) JP-A-63-313035 (JP, A) JP-A-2-105042 (JP, A) JP-A-63-109370 (JP) , A)

Claims (4)

(57) [Claims] 1. An electrophoresis pattern reading apparatus which excites a fluorescent substance labeling a sample after electrophoresis and development of the sample to emit fluorescence, and reads a fluorescent pattern to emit light, wherein the fluorescent substance of the electrophoretic pattern is A light source that emits irradiation light to be excited, a light scanning mechanism that scans irradiation light from the light source along a predetermined optical axis and irradiates the gel in the thickness direction, and a light receiving surface in a direction different from the optical axis of the irradiation light. A light-receiving unit that separates and receives the fluorescence of the migration pattern from the scattered light on the reading surface according to the spatial positional relationship of the light-receiving path, and a photoelectric conversion that photoelectrically converts the optical signal received by the light-receiving unit and outputs an electric signal Unit, an amplifier that performs an integration operation corresponding to the scanning of the irradiation light with respect to the electric signal from the photoelectric conversion unit, amplifies and sequentially outputs a fluorescent electric signal from the migration pattern, and an electric signal from the amplifier. axis An optical axis correction unit that receives the detection signal and corrects the optical axis of the irradiation light. 2. The optical axis correction unit includes: a folding mirror that changes an optical axis of irradiation light from a light source; an actuator for correcting an optical axis that changes an angle of the folding mirror; an actuator driver that drives the actuator; 2. The fluorescent pattern reading device according to claim 1, further comprising an optical axis correction mechanism including a control circuit for providing a drive control signal to an actuator driver. 3. The actuator for optical axis correction is a ceramic actuator, and the control circuit receives an electric signal from an amplifier as an optical axis detection signal, and the control circuit receives the electric signal from the amplifier in a direction in which the magnitude of the optical axis detection signal becomes an extreme value. 3. The fluorescence pattern reading device according to claim 2, wherein the device is a feedback control circuit that generates a drive control signal to an actuator driver. 4. The optical axis correcting actuator comprises a plurality of ceramic actuators, and the plurality of ceramic actuators constitute a drive system for displacing an optical axis reflection angle of one folding mirror. The fluorescence pattern reading device according to claim 2.