JP2004333333A - Dna microarray substrate and reading device - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a system having a simple constitution, extremely high efficiency and a low price, capable of reexamining fundamentally the structure of a DNA microarray or a scanner or the like, reading a DNA spot stably in a short time, and acquiring an analysis result. <P>SOLUTION: This device for reading the DNA microarray formed by arranging the DNA spot on a substrate is equipped with an optical system for irradiating the DNA microarray with laser light having the wavelength λ1 and exciting a phosphor included in the DNA spot by the laser light, an objective lens for condensing excited fluorescence having the wavelength λ2, the first photodetector for converting the fluorescence having the wavelength λ2 condensed by the objective lens into an electric signal, and a modulation means for modulating the amplitude of output light of the laser light. The reading device characterized by reading the DNA spot on the DNA microarray, or the like is provided. <P>COPYRIGHT: (C)2005,JPO&NCIPI

Description

[0001]
TECHNICAL FIELD OF THE INVENTION
The present invention relates to a configuration of a DNA microarray substrate and a measuring device.
[0002]
[Prior art]
A DNA microarray is one in which thousands of DNA probes are immobilized on a substrate such as a slide glass. A sample (target) labeled with a fluorescent molecule or the like is allowed to flow, and is hybridized with the probe. Is measured to estimate the expression level of the gene contained in the sample.
As a result, the expression of a large number of genes can be analyzed comprehensively at the same time, so that it is widely used as a standard instrument for research and development in the fields of life science, pharmacy, and agriculture.
For example, mRNA (messenger RNA) is extracted from a cancer tissue that responds to a drug and a cancer tissue that does not react, and the difference between the expression levels of the two is estimated by a DNA microarray carrying all human genes. Specific gene expression is found. Using this, the onset mechanism can be elucidated and drug discovery research can be efficiently carried out targeting the gene.
[0003]
In the era of post-genome sequencing, the need for comprehensive measurement of gene expression has spread to the new biotechnology industry, such as food inspection and production quality control, as well as clinical testing in the so-called tailor-made medicine and medical fields. The market is expected to grow significantly.
There are two main types: the Affymetrix type, in which oligonucleotides are vertically stacked on a silicon substrate by photolithography and solid-phase DNA synthesis, and the so-called Stanford type, in which DNA is attached to a slide glass. is there. The former method is expensive because it requires selecting a gene in advance and requesting design and manufacture, while the latter method has the advantage that the user can freely select the gene to be spotted in the use environment.
[0004]
[Problems to be solved by the invention]
In the current DNA microarray, the fluorescence of DNA spots arranged in a two-dimensional lattice is imaged and read by laser scanning or image measurement means. The measurement requires two degrees of freedom, which complicates the apparatus, and also requires post-processing such as correction of a fluorescence signal and computer image analysis processing for noise reduction.
Although there are devices using beads instead of slide glasses and devices using porous media, the coverage is limited to at most about 100 types of genes.
[0005]
However, systems consisting of spotters, hybridizations, and scanners for analyzing DNA microarrays are challenging in terms of quantification and sensitivity.
If you list your current issues,
1. Poor fluorescence measurement sensitivity
2. It takes time to read
3. Imaging by two-dimensional scanning is required
4. Poor operability
5. Not enough DNA spots to be placed on one glass substrate (at most 10,000 spots)
6. It was difficult to record the properties of the samples and the experimental conditions at the same time, so the samples were numbered and data sheets were used separately.
There were issues such as.
[0006]
It is an object of the present invention to fundamentally review the structure of a DNA microarray or a scanner, stably read a DNA spot in a short time, obtain an analysis result, and have a simple configuration, a very efficient and low-cost system. Is to provide.
The spots of the DNA microarray currently arranged in a two-dimensional X, Y two-dimensional lattice are arranged one-dimensionally, and at the same time, the glass plate is changed to a disk shape, and an index capable of specifying the spot position is provided. We propose a new optical system and servo system to improve the detection S / N of the spot scanner, and realize high-speed measurement, improved operability, and low cost.
[0007]
[Means for Solving the Problems]
The present invention relates to, for example, the following inventions.
A. In an apparatus for reading a DNA microarray in which DNA spots are arranged on a substrate,
An optical system for irradiating the DNA microarray with laser light having a wavelength of λ1 so that the laser light excites a phosphor contained in the DNA spot;
An objective lens for condensing the fluorescence of the excited wavelength λ2,
A first photodetector for converting the fluorescence of wavelength λ2 collected by the objective lens into an electric signal;
Comprising a modulating means for amplitude modulating the output light of the laser light,
A reading device for reading DNA spots on the DNA microarray.
[0008]
B. In an apparatus for reading a DNA microarray in which DNA spots are arranged on a substrate,
An optical system that irradiates the DNA microarray with laser light having a wavelength of λ1 so that the laser light excites a phosphor contained in the DNA spot;
An objective lens for condensing the fluorescence of the excited wavelength λ2,
A first photodetector for converting the fluorescence of wavelength λ2 collected by the objective lens into an electric signal;
A second photodetector for mainly detecting reflected light of wavelength λ1 reflected by the irradiated laser light of wavelength λ1 from the substrate on which the DNA spot is arranged;
Position detection means for detecting the vertical relative position of the objective lens with respect to the substrate, using an output of the second photodetector as an input,
An actuator for displacing the objective lens in a direction perpendicular to the substrate,
A drive device for displacing the output of the position detection means in a vertical direction of the actuator of the objective lens,
A reading device for reading DNA spots on the DNA microarray.
[0009]
C. In an apparatus for reading a DNA microarray in which DNA spots are arranged on a substrate,
An optical system that irradiates the DNA microarray with laser light having a wavelength of λ1 so that the laser light excites a phosphor contained in the DNA spot;
An objective lens for condensing the fluorescence of the excited wavelength λ2,
A first photodetector for converting the fluorescence of wavelength λ2 collected by the objective lens into an electric signal;
A second photodetector for mainly detecting reflected light of wavelength λ1 reflected by the irradiated laser light of wavelength λ1 from the substrate on which the DNA spot is arranged;
Position detection means for detecting a vertical relative position of the objective lens with respect to the substrate using an output of the second photodetector;
An actuator for displacing the objective lens in a direction perpendicular to the substrate,
A drive device for displacing the output of the position detection means in a vertical direction of the actuator of the objective lens,
The reflected light of the wavelength λ1 is detected by at least a two-divided photodetector, and the position of the objective lens in the X direction that is parallel to the substrate is controlled by a difference between outputs of the two-divided photodetectors, A control device for causing the laser light to follow the DNA spot train,
Comprising a modulating means for amplitude modulating the output light of the laser light,
A reading device for reading DNA spots on the DNA microarray.
[0010]
D. In an apparatus for reading a DNA microarray in which DNA spots are arranged on a substrate,
An optical system that irradiates the DNA microarray with laser light having a wavelength of λ1 so that the laser light excites a phosphor contained in the spot;
An objective lens for condensing the fluorescence of the excited wavelength λ2,
A first photodetector for converting the fluorescence of wavelength λ2 collected by the objective lens into an electric signal;
A second photodetector for mainly detecting reflected light of wavelength λ1 reflected by the irradiated laser light of wavelength λ1 from the substrate on which the spot is arranged;
Position detection means for detecting, as an input, an output of the second photodetector, a relative position Z of the objective lens in a vertical direction with respect to the substrate;
An actuator for displacing the objective lens in a direction perpendicular to the substrate,
A signal generator for displacing the objective lens in a vertical direction, a driving device having an output of the signal generator as a first input,
The drive unit for displacing the actuator of the objective lens in the Z direction, wherein the output of the position detection unit is a second input;
Phase-detecting the signal displaced in the Z direction and the output of the second photodetector, inputting the output of the phase detector to the driving device,
A reading device for reading spots on the DNA microarray.
[0011]
E. FIG. In an apparatus for reading a DNA microarray in which DNA spots are arranged on a substrate,
An optical system that irradiates the DNA microarray with laser light having a wavelength of λ1 so that the laser light excites a phosphor contained in the spot;
An objective lens for condensing the fluorescence of the excited wavelength λ2,
A first photodetector for converting the fluorescence of wavelength λ2 collected by the objective lens into an electric signal;
A second photodetector for mainly detecting reflected light of wavelength λ1 reflected by the irradiated laser light of wavelength λ1 from the substrate on which the spot is arranged;
Position detection means for detecting the vertical relative position of the objective lens with respect to the substrate, using an output of the second photodetector as an input,
An actuator for displacing the objective lens in a direction perpendicular to the DNA spot row on the substrate and in an X direction parallel to the substrate;
A signal generator and a driving device for displacing the objective lens in the X direction;
Phase-detecting the signal displaced in the X direction and the output of the second photodetector, and inputting the output of the phase detector to the actuator driving device;
A reading device for reading spots on the DNA microarray.
[0012]
BEST MODE FOR CARRYING OUT THE INVENTION
In the present invention, an optical system for improving the detection S / N of a spot scanner and a servo system are proposed to realize high-speed measurement, improved operability, and reduced cost. An optical system constituting the servo system is provided independently of the optical system for reading the DNA spot, so that the beam spot for detecting the servo error does not deteriorate the phosphor contained in the DNA spot. In addition, the position of the reading light beam of the DNA microarray is accurately controlled by using a servo mechanism to eliminate errors in the focusing and tracking directions and to perform accurate scanning.
[0013]
Further, the spots of the DNA microarray currently arranged in a two-dimensional lattice are arranged concentrically or spirally in one dimension, and at the same time, the glass plate is changed to a disk shape, and an index capable of specifying the spot position is provided. Specifically, spotting is accurately performed at a position on the glass disk where an index is provided, and scanning is performed in a one-dimensional direction to read a DNA spot without distortion.
Further, a recordable zone is provided on the DNA microarray, and preparation conditions and measurement results are recorded on the target DNA microarray. As a result, the glass plate and the data at the time of creation, which have been separately stored, can be integrally stored, thereby improving security.
Further, in order to prevent fading of the phosphor by the reading laser beam and to improve the reading S / N of the DNA spot, the reading laser beam is amplitude-modulated at a high frequency.
[0014]
【Example】
DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS An embodiment specifically showing the best mode for carrying out the present invention will be described below with reference to the drawings.
Hereinafter, a schematic configuration of the first embodiment of the present invention will be described with reference to FIGS. 1, 2, and 3. FIG.
[0015]
FIG. 1 is a block diagram showing a configuration of an apparatus for reading the DNA microdisk (abbreviated as DNA-MD) shown in FIG. 2, and FIG. 2 is a DNA microdisk.
In FIG. 1, reference numeral 1 denotes a DNA micro disk, and its rotation is controlled by a disk motor 2. Reference numeral 3 denotes an objective lens which focuses a laser beam on a DNA spot formed on a DNA microdisk and collects reflected light. The 3rd objective lens has a 4th objective lens actuator and a focusing element 4a and a 4th tracking element 4b, so that the laser beam can follow the direction perpendicular to the substrate of the DNA micro disk (Z direction of 3b) and the DNA spot array. In the direction (X direction of 3a).
[0016]
Reference numeral 5 denotes a fluorescence excitation light source (output light wavelength λ1), in which case a laser having a wavelength of 650 nm is used. Reference numeral 6 denotes a collimator lens 1 which converts the output light of the laser 5 into a parallel light or a divergent light having a prescribed angle, passes through a half mirror 9, a beam splitter 1, and an objective lens 3. Focus on one DNA microdisk. The reason why the light is converted into divergent light having a specified angle is to make the size of the beam spot at the time of focusing equal to the size of the DNA spot. At this time, the light beam emitted from the light source having the wavelength λ1 is provided with an aperture limit before entering the objective lens, so that the NA of the objective lens is substantially reduced. This aperture restriction can be provided on the output side of the collimator lens 8 in FIG. For example, even when the objective lens has a NA of 6.0, the NA is 5.0 for light of wavelength λ1, and 6.0 for servo light of wavelength λ3. I did it. By doing so, the servo error detection sensitivity can be increased, and the diameter of the DNA spot reading beam can be increased.
Reference numeral 7 denotes a servo light source (output light wavelength λ3), which passes through a collimator lens 2, a half mirror 9, a beam splitter 1, a beam splitter 1, and an objective lens 3 to focus on one DNA microdisk. Tie. The light reflected by the DNA microdisk passes through the objective lens 3, the beam splitter 1, the beam splitter 2, the condensing lens 2, and the servo error detector 16. .
[0017]
On the other hand, the output light of the fluorescence excitation light source (output light wavelength λ1) collected on one DNA microdisk by the three objective lenses is a fluorescent substance contained in a DNA spot on the one DNA microdisk. The excited and excited light having the wavelength λ2 is collected by the objective lens 3, passes through the beam splitters 10 and 14, is condensed by the condensing lens 1, and is then excited by the optical filter 12. Only the wavelength of the light is selected and directed to thirteen fluorescence reading detectors. At this time, the beam splitters 10 and 14 pass the emission wavelength λ2 of the fluorescent substance in the DNA spot excited by the laser having the wavelength λ1, and the beam splitter 14 emits the light having the output wavelength λ3 of the servo light source. The twelve optical filters are configured to reflect and transmit only λ2. The output light from the servo light source 7 is converted into parallel light by the collimator lens 2.
[0018]
When the spot is condensed by the three objective lenses by the seventeen cylindrical lenses, the shape of the condensed spot changes before and after the focal point. In other words, the circle changes from a perfect circle to an ellipse on the 16 servo error detectors. This change is measured by a servo error detector obtained by dividing 16 photodetectors into four. When the beam reflected from the DNA microdisk is projected in a substantially circular state at the center of the detector, it indicates that the three objective lenses are in focus. At the same time, the output light of the servo light source is converted into light beams S1, S2, S3 emitted from the objective lens 3 on a DNA spot or a pre-group on one DNA microdisk by a diffraction grating as shown in FIG. Is formed. Of these, the light beams indicated by S1 and S3 are received by D1 and D2 of 16 servo error detectors, and the difference between their outputs becomes a tracking error.
[0019]
In FIG. 2, reference numeral 21 denotes a substrate, which is made of glass or resin. Reference numeral 22 denotes a center hole provided with a hole at the center of the substrate. Reference numeral 23 denotes a DNA spot formed on the substrate in a spiral or concentric manner. The term “DNA spot” as used herein means a droplet containing a probe DNA (general term including DNA spots after hybridization and referred to as DNA spots). Here, the diameter of the DNA spot is set to 250 μm or less.
Reference numeral 24 denotes a first data recording unit, which prepares and records a row of spots modulated by the information such as conditions for forming a DNA spot when the DNA spot is provided on the substrate. As a modulation method that can be used at this time, a method of changing the position of the spot according to an information signal composed of a binary signal, a method of changing the period of the spot according to the information signal, or the like is used. It is possible. In addition, as a material forming the spot, an ink made of an organic material, an inorganic material, or the like can be used.
[0020]
Although not shown, the data of the DNA spot and the data of 24 can also be arranged on a pre-group provided on the 21 substrates.
As a method of creating a pre-groove, a substrate can be selectively etched and created. In addition, the convex portion of the pre-groove can be provided by printing, and when the pre-groove is formed by printing, the DNA spot is arranged on the pre-groove to which the printing ink does not adhere.
A substrate provided with DNA spots on such a substrate is called a DNA microdisk. After the DNA spot was provided on the DNA microdisk, the DNA spot was inserted into the cartridge, and then the DNA microdisk was read from the reading window provided on the cartridge.
[0021]
In addition, the data recording portion is provided by modulating the DNA spot row when creating the DNA spot, but a second recordable portion 25 is provided on the substrate separately from the recording portion, and after reading the DNA spot, It is also possible to additionally write information data of the read DNA spot.
In the recordable portion, a dye or a metal thin film conventionally used for a recordable optical disk can be formed on a substrate by vapor deposition or sputtering. It is preferable that the second data recording section is provided on the pre-groove, and that the pre-groove is provided with address information to facilitate identification with other parts. This recording method will be described later with reference to FIG. The above-mentioned first and second recording units can be added to the following conventional DNA microarray substrate.
[0022]
FIG. 3 shows a conventionally used DNA microarray in which DNA spot rows 25 are arranged on a substrate 26 in a two-dimensional lattice. Numeral 27 indicates the direction in which the DNA spot is scanned.
The reading device shown in FIG. 1 is configured to be able to read any of the DNA microdisk and the DNA microarray shown in FIGS. When reading the DNA microarray, a mechanism (not shown) for moving the DNA microarray in parallel is used instead of the two disk motors.
[0023]
FIG. 4 is a principle diagram showing a relative relationship between a beam spot formed by a DNA microdisk, a reading laser beam, and a servo laser beam.
In FIG. 4, reference numeral 31 denotes a pregroup, which is indicated by P1, P2, and P3. The width of the pre-group is set to 5 to 100 μm, the height is set to a convex or concave of 3 to 10 μm, and the interval of the pre-group is set to about 5 to 150 μm. 34 DNA spots are formed on 31 pregrooves. Reference numeral 33 denotes a servo beam, which indicates a state where 31 pregrooves P2 are irradiated.
The servo beam S1 irradiates the servo error detector D1 shown in FIG. 1, S2 irradiates D3, and S3 irradiates D2.
Reference numeral 32 denotes a DNA spot reading beam (wavelength λ1), which irradiates the DNA spot of 34 and excites the phosphor contained in the DNA spot of 34, and the excitation light of wavelength λ2 by the wavelength λ1 is the fluorescence reading detection light of FIG. It is read by the device 13. The servo beam formed from the servo light source (wavelength λ3) irradiates the DNA spot, but it is set so as not to have a wavelength that can excite the phosphor of the DNA spot (for example, 780 nm). Absent.
[0024]
FIG. 5 is a block diagram showing a control system and a signal measurement system of the reading device shown in FIG. The overall configuration is the same as that of the reading apparatus shown in FIG. 1, and the same components are represented by the same numbers.
In FIG. 5, reference numeral 16 denotes a servo error detector, and D3 denotes a quadrant photodetector for detecting a focus error. The light output of the diagonal photodetector of D3 is converted into an electric signal by the adders 41 and 45. A difference between the outputs of the adders 41 and 45 is detected by a differential amplifier 42 to obtain a focus error signal. This principle will be described with reference to the principle diagram of the focus error signal in FIG.
The output of the differential amplifier 42 is adjusted in servo response by the phase compensator 43, and then the focus amplifier 4b is driven by the drive amplifier 44 to control the position of the objective lens 3 in the vertical direction. .
Further, the outputs of the photodetectors D1 and D2 are input to the differential amplifier 46, and a tracking error indicating the position of the pre-groove signal or the DNA spot train is detected from the differential amplifier output 46.
The output of the differential amplifier 46 passes through the phase compensator 47 and is input to the drive amplifier 48 to control the position of the tracking element 4a.
A CPU (Central Processing Unit) 49 performs control operations such as pull-in, monitoring, and sequence control of the servo system, and controls the entire servo system.
[0025]
FIG. 6 is a diagram showing the correlation between the focus error and the position of the objective lens.
In FIG. 6, a photodetector D3 of the servo error detector is shown, and a distribution of light projected on the photodetector D3 is shown. Reference numeral 52 denotes a focused state, the light distribution on the photodetector D3 is substantially circular, and the output of the differential amplifier 42 in FIG.
Reference numeral 51 indicates a state in which the objective lens is close to the substrate of the DNA microdisk. At this time, the output of the differential amplifier 42 outputs a negative voltage, and 53 indicates a state in which the objective lens is far away, and a positive voltage is output. In this way, it is possible to apply a voltage to the focus element 4b and control the drive so that the distance between the objective lens 3 and the DNA microdisk 1 is in focus.
Further, it is possible to reduce the fading of the phosphor by modulating the fluorescence excitation light source at a frequency of, for example, about 400 MHz and irradiating the modulated light to the DNA spot on the DNA microdisk. I understand.
By doing so, the duty of color development of the phosphor can be reduced. Further, by modulating the laser of the light source, it is possible to suppress the generation of noise when return light enters the laser.
[0026]
FIG. 8 is a block diagram of a data recording system for additionally writing data obtained by measuring characteristics of a DNA spot on the DNA microdisk on the DNA microdisk of FIG. 7.
The overall configuration is the same as that of the reading device shown in FIGS. 1 and 5, and the components are represented using the same numbers.
The data recording is for irradiating the recording film of the second data recording section 25 provided on one DNA micro disk with a recording laser beam modulated by data to perform recording. In FIG. 7, reference numeral 64 denotes a personal computer (PC) which prepares data obtained by measuring characteristics of a DNA spot to be recorded, encodes the data to be recorded by an encoder of 62 via an interface 63, and a modulator of 61 Thus, the amplitude of the output light of the servo light source or the fluorescence excitation light source 7 is modulated, and the recording film is irradiated with the modulated laser light to perform the recording operation. In order to accurately detect the recording section, the position of the pre-groove provided with the recording section is determined by reading the address information.
[0027]
The recording film undergoes a physical change (a hole or deformation in the film) due to the modulated laser light, and data is recorded. In order to reproduce the recorded data, the power of the output light of the servo light source 7 is set to a low power for reading, and the reflected light from the recording film is used using the output of the 16 servo error detector. The outputs of the adders 41 and 45 are supplied to an adder 66, and an output signal of the adder is demodulated by a decoder 67. As an encoding method, for example, PPM (pit position modulation) expressing data by the position of a recording spot is used.
When the recorded data is read, the data demodulated by the decoder 67 is sent to the PC 64 via the interface 63 and error-corrected, and the data is read.
[0028]
FIG. 8 is a block diagram of a wobbling servo system showing a tracking servo system for causing a read beam to follow a DNA spot row. The overall configuration is the same as that of the reading device shown in FIGS. 1 and 5, and the components are represented using the same numbers.
An adder 71 adds the outputs of the servo error detector. It should be noted that only the photodetector D3 is used among the servo error detectors. Reference numeral 72 denotes an oscillator, which is input to a driving amplifier 48 and vibrates the tracking element 4a minutely (about 1 μm). A filter 74 receives the output of the adder 71, extracts only the frequency component of the oscillator 72, and inputs it to the phase detector 73. The output of the phase detector indicates a tracking error, and the drive element 48 controls the tracking element 4a constituting the objective lens actuator 4 to follow the DNA spot array on the DNA microdisk.
When a DNA microarray is used, the position of the objective lens is controlled such that a laser beam applied to the DNA spot array on the DNA microarray from the objective lens follows the DNA spot array.
[0029]
FIG. 9 shows a second embodiment in which the reader shown in FIG. 1 is partially modified. The overall configuration is the same as that of the reading apparatus shown in FIG. 1, and the same components are represented by the same numbers. The difference from the embodiment shown in FIG. 1 is that the tracking error signal is detected using a phosphor excitation light source. The diffraction grating 18 used in FIG. 1 and the photodetectors D1 and D2 of the servo error detector are not used. Instead, a beam splitter 3 at 81, a condenser lens at 82, and a two-segment photodetector 83 are added.
In FIG. 9, the light of wavelength λ1 emitted from the light source for fluorescence excitation 5 is reflected by the beam splitter 10 and irradiates the DNA microdisk through the objective lens 3 and the reflected light is the beams 10 and 14. The light passes through the splitter, is reflected by the beam splitter 81, and reaches the split photodetector 83 by the condenser lens 82. A DNA spot array or a pregroove on the DNA microdisk is projected onto the 83-divided photodetector 83, and the difference between the respective photodetectors of the 83-divided photodetector is calculated to obtain a DNA spot array or a pregroove. Is obtained. This technique is disclosed in, for example, Japanese Patent Registration No. 2134883 which is a patent of the inventor. By using the difference as the tracking error shown in FIG. 5 and controlling the position of the objective lens, a light beam of wavelength λ1 emitted from the fluorescence excitation light source 5 is irradiated onto the DNA spot of one DNA microdisk. be able to. In the present embodiment, the wavelength of the reading beam is set to one type of λ1, but of course, it is also possible to add a light source having a different wavelength and simultaneously irradiate the DNA spot to read the DNA spot. In that case, one of the plurality of read beams can be a servo beam.
In order to make the beam for reading the DNA spot approximately the same as the size of the DNA spot on the DNA microarray, an aperture limit is inserted in the light emitted from the collimator lens 6 and only the light passing near the center of the objective lens 3 is used. Should focus on the DNA spot. On the other hand, for the servo beam of wavelength λ3, in order to increase the sensitivity of detecting the position of the objective lens in the servo error signal, it is better not to limit the aperture of the objective lens or to set it smaller than the aperture limit of the reading beam. preferable.
[0030]
With the above-described configuration, it is possible to detect the position of the DNA spot array and control the position of the objective lens using the light source for fluorescence excitation, so that a highly accurate tracking servo can be realized.
[0031]
【The invention's effect】
The present invention configured as described above has the following effects, and can solve the problem. A laser beam for irradiating a DNA spot on a DNA microdisk or a DNA microarray has both a servo laser beam and a reading laser beam, so that the reading laser beam can be accurately positioned on the DNA spot, and the DNA spot can be positioned. Was efficiently excited, and the sensitivity of the fluorescence measurement was improved. Of course, even if there is a deformation such as warpage of the substrate on which the DNA microspot is placed, or if the placement position of the DNA spot is displaced, the accurate position of the spot can be detected by the servo and the reading beam can accurately irradiate the spot. Was. Further, by modulating the read beam at a high frequency, the fading of the phosphor is prevented, and a higher read laser peak power than before can be applied.
In particular, when a microdisk substrate is used, the reading beam can be scanned in the one-dimensional direction, so that the reading speed can be increased. Therefore, the scanning is completed only once when the DNA spot is scanned once, and the imaging by the conventional two-dimensional scanning is not required.
Further, since the DNA spot was provided on the DNA microdisk and then inserted into the cartridge, handling was improved.
Furthermore, since a DNA microdisk having a pregroove provided on the substrate is used, more DNA spots can be provided on the substrate than before, and a DNA microdisk having, for example, 100,000 or more DNA spots can be manufactured.
[Brief description of the drawings]
FIG. 1 is a block diagram illustrating a schematic configuration of a reading apparatus according to an embodiment of the invention.
FIG. 2 is a diagram showing a schematic configuration of a DNA microdisk of the present invention.
FIG. 3 is a diagram showing a schematic configuration of a conventional DNA microarray.
FIG. 4 is a diagram showing a relative positional relationship between a read beam and a servo beam on a DNA microdisk.
FIG. 5 is a block diagram of a reading device servo system.
FIG. 6 is a principle diagram showing a relationship between a focus error and a position of an objective lens.
FIG. 7 is a block diagram of a data recording system.
FIG. 8 is a block diagram showing tracking control of a wobbling method.
FIG. 9 is a second embodiment of the reading device.
[Explanation of symbols]
1 DNA microdisk
2 Disk motor
3 Objective lens
4. Objective lens actuator (4a tracking element, 4b focus element)
5 Light source for fluorescence excitation (output light wavelength λ1)
6 Collimator lens 1
7. Light source for servo (output light wavelength λ3)
8 Collimator lens 2
9 Half mirror
10 Beam splitter 1
11 Condensing lens 1
12 Optical filter
13 Fluorescence reading detector
14 Beam splitter 2
15 Condensing lens 2
16 Servo error detector
16 Second photodetector
17 Cylindrical lens
18 diffraction grating
21 Substrate
22 center hole
23 DNA spots
24 First Data Recording Unit
25 Second data recording unit
26 substrate
27 Scanning direction
31 pre-groups
32 DNA spot read beam (wavelength λ1)
33 Servo beam
34 DNA spots
41 Adder
42 differential amplifier
43 Phase compensator
44 Drive amplifier
45 Adder
46 Differential amplifier
47 Phase compensator
48 drive amplifier
49 CPU (Central Processing Unit)
61 Modulator
62 encoder
63 Interface
64 Personal computer (PC)
66 adder
67 decoder
71 Adder
72 oscillator
73 phase detector
81 Beam Splitter 3
82 Condensing lens 3
83 split photodetector

Claims (19)

In an apparatus for reading a DNA microarray in which DNA spots are arranged on a substrate,
An optical system for irradiating the DNA microarray with laser light having a wavelength of λ1 so that the laser light excites a phosphor contained in the DNA spot;
An objective lens for condensing the fluorescence of the excited wavelength λ2,
A first photodetector for converting the fluorescence of wavelength λ2 collected by the objective lens into an electric signal;
Comprising a modulating means for amplitude modulating the output light of the laser light,
A reading device for reading DNA spots on the DNA microarray. In an apparatus for reading a DNA microarray in which DNA spots are arranged on a substrate,
An optical system that irradiates the DNA microarray with laser light having a wavelength of λ1 so that the laser light excites a phosphor contained in the DNA spot;
An objective lens for condensing the fluorescence of the excited wavelength λ2,
A first photodetector for converting the fluorescence of wavelength λ2 collected by the objective lens into an electric signal;
A second photodetector for mainly detecting reflected light of wavelength λ1 reflected by the irradiated laser light of wavelength λ1 from the substrate on which the DNA spot is arranged;
Position detection means for detecting the vertical relative position of the objective lens with respect to the substrate, using an output of the second photodetector as an input,
An actuator for displacing the objective lens in a direction perpendicular to the substrate,
A drive device for displacing the output of the position detection means in a vertical direction of the actuator of the objective lens,
A reading device for reading DNA spots on the DNA microarray. In an apparatus for reading a DNA microarray in which DNA spots are arranged on a substrate,
An optical system that irradiates the DNA microarray with laser light having a wavelength of λ1 so that the laser light excites a phosphor contained in the DNA spot;
An objective lens for condensing the fluorescence of the excited wavelength λ2,
A first photodetector for converting the fluorescence of wavelength λ2 collected by the objective lens into an electric signal;
A second photodetector for mainly detecting reflected light of wavelength λ1 reflected by the irradiated laser light of wavelength λ1 from the substrate on which the DNA spot is arranged;
Position detection means for detecting a vertical relative position of the objective lens with respect to the substrate using an output of the second photodetector;
An actuator for displacing the objective lens in a direction perpendicular to the substrate,
A drive device for displacing the output of the position detection means in a vertical direction of the actuator of the objective lens,
The reflected light of the wavelength λ1 is detected by at least a two-divided photodetector, and the position of the objective lens in the X direction that is parallel to the substrate is controlled by a difference between outputs of the two-divided photodetectors, A control device for causing the laser light to follow the DNA spot train,
A reading device for reading DNA spots on the DNA microarray. In order to detect the distance between the objective lens and the substrate, a light source having a wavelength λ3 used for detecting a servo signal for irradiating the substrate is provided independently of a light source having a wavelength λ1 for exciting a phosphor of a DNA spot, and the servo 3. The reader according to claim 2, wherein the wavelength λ3 of the signal detection light source is set to a wavelength that does not excite the phosphor. In order to detect the distance between the objective lens and the substrate, a light source having a wavelength λ3 used for detecting a servo signal for irradiating the substrate is provided independently of a light source having a wavelength λ1 for exciting a phosphor of a DNA spot; The wavelength λ3 of the signal detection light source is selected to a wavelength that does not excite the phosphor,
The aperture of the light beam having the wavelength λ1 for irradiating the DNA spot incident on the objective lens is substantially limited, and the servo signal detection light source has a beam diameter larger than that of the beam formed on the substrate. 3. The reading device according to claim 2, wherein: In order to detect the distance between the objective lens and the substrate, a light source having a wavelength λ3 for detecting a servo signal for irradiating the substrate is provided independently of a light source for exciting a phosphor of a DNA spot, and While selecting the wavelength of the light source to a wavelength that does not excite the phosphor,
Light output from a light source for exciting the fluorescent material of the DNA spot is reflected by the substrate, and the reflected light of wavelength λ1 is used to detect the position of the DNA spot in the X direction and irradiate the DNA spot. 3. The reader according to claim 2, wherein the position of the objective lens in the X direction is controlled so that the light follows the DNA spot. In order to detect the distance between the objective lens and the substrate, a servo signal detection light source for irradiating the substrate is provided independently of a light source for exciting a DNA spot fluorescent substance, and a wavelength λ3 of the servo signal detection light source is provided. Is selected so as not to excite the phosphor,
A servo error detector that detects a vertical position of the substrate on which the DNA spot is disposed, using light reflected from the substrate, the light output from the light source having the wavelength λ3 for detecting the servo signal;
3. The reader according to claim 2, wherein the position of the objective lens is controlled such that a light beam having a wavelength λ1 irradiating the DNA spot follows the DNA spot. In order to detect the distance between the objective lens and the substrate, a servo signal detection light source for irradiating the substrate is provided independently of a light source for exciting a DNA spot fluorescent substance, and a wavelength λ3 of the servo signal detection light source is provided. Is selected so as not to excite the phosphor,
A servo error detector that detects a vertical position of the substrate on which the DNA spot is disposed, using light reflected from the substrate, the light output from the light source having the wavelength λ3 for detecting the servo signal;
A control system for controlling the position of the objective lens so that the light beam of wavelength λ1 irradiating the DNA spot follows the DNA spot,
3. The reader according to claim 2, wherein the light beam for detecting the servo error is set so as not to irradiate the DNA spot. In order to detect the distance between the objective lens and the substrate, a servo signal detection light source for irradiating the substrate is provided independently of a light source for exciting a phosphor of a DNA spot, and the servo signal detection light source is Select a wavelength that does not excite when illuminating the phosphor,
A diffraction grating is inserted into an output optical path of the servo light source, and at least the output light forms three beam spots on a DNA spot row on the substrate,
The reflected light of the two beam spots among the three beam spots is detected by a photodetector, the output of the photodetector is input to a differential amplifier, and the difference signal indicates the position of the DNA spot row. Then, the position of the objective lens in the X direction perpendicular to the DNA spot row for irradiating the beam spot to the substrate and parallel to the substrate is set so that the difference signal indicating the position becomes a predetermined value. A DNA spot reader which controls movement. A pre-groove is provided on the substrate, and a droplet containing probe DNA is arranged in a convex portion or a concave portion of the pre-groove. The spread of the pre-groove in a direction perpendicular to the groove is restricted by the surface tension of the droplet. A DNA microarray substrate, which extends in a tangential direction and in which probe DNA is bound on the substrate. 11. The DNA microarray substrate according to claim 10, wherein a pregroove is provided on the substrate, and address information of the pregroove is provided on a convex portion or a concave portion of the pregroove. 12. The DNA microarray substrate according to claim 11, wherein DNA spots are arranged on the substrate in a spiral or concentric manner. When a DNA spot is formed on a substrate, information such as conditions for forming the DNA spot is provided, and an area for forming and recording a row of spots modulated by the information is provided. Microarray substrate. A DNA microarray substrate, comprising: a recording section provided with a recording film capable of recording data by irradiating a laser beam onto a region where the DNA spot is not arranged. A DNA microarray substrate, wherein the pregroove-shaped irregularities on the DNA microarray substrate have a width of 5 μm or more and 200 μm or less. A DNA microarray substrate, wherein the irregularities of the pregroove are formed by printing. A DNA microarray substrate, wherein a length of a tangential direction on a pregroove of a region where the probe DNA is provided on the DNA microarray substrate is twice or more as compared with a width in a direction perpendicular to the pregroove. In an apparatus for reading a DNA microarray in which DNA spots are arranged on a substrate,
An optical system that irradiates the DNA microarray with laser light having a wavelength of λ1 so that the laser light excites a phosphor contained in the spot;
An objective lens for condensing the fluorescence of the excited wavelength λ2,
A first photodetector for converting the fluorescence of wavelength λ2 collected by the objective lens into an electric signal;
A second photodetector for mainly detecting reflected light of wavelength λ1 reflected by the irradiated laser light of wavelength λ1 from the substrate on which the spot is arranged;
Position detection means for detecting, as an input, an output of the second photodetector, a relative position Z of the objective lens in a vertical direction with respect to the substrate;
An actuator for displacing the objective lens in a direction perpendicular to the substrate,
A signal generator for displacing the objective lens in a vertical direction, a driving device having an output of the signal generator as a first input,
The drive unit for displacing the actuator of the objective lens in the Z direction, wherein the output of the position detection unit is a second input;
Phase-detecting the signal displaced in the Z direction and the output of the second photodetector, inputting the output of the phase detector to the driving device,
A reading device for reading spots on the DNA microarray. In an apparatus for reading a DNA microarray in which DNA spots are arranged on a substrate,
An optical system that irradiates the DNA microarray with laser light having a wavelength of λ1 so that the laser light excites a phosphor contained in the spot;
An objective lens for condensing the fluorescence of the excited wavelength λ2,
A first photodetector for converting the fluorescence of wavelength λ2 collected by the objective lens into an electric signal;
A second photodetector for mainly detecting reflected light of wavelength λ1 reflected by the irradiated laser light of wavelength λ1 from the substrate on which the spot is arranged;
Position detection means for detecting the vertical relative position of the objective lens with respect to the substrate, using an output of the second photodetector as an input,
An actuator for displacing the objective lens in a direction perpendicular to the DNA spot row on the substrate and in an X direction parallel to the substrate;
A signal generator and a driving device for displacing the objective lens in the X direction;
Phase-detecting the signal displaced in the X direction and the output of the second photodetector, and inputting the output of the phase detector to the actuator driving device;
A reading device for reading spots on the DNA microarray.

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