JP2009229378A - Capillary array sheet inspection apparatus and inspection method - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a capillary array sheet inspection apparatus and inspection method capable of positively and rapidly detecting a defect of gel of a through-hole of the capillary array sheet. <P>SOLUTION: The capillary array sheet inspection apparatus is equipped with an illuminating device 14 equipped with a light source 12, a grid pattern 22 irradiated with light from the light source, a capillary array sheet holding means 24 for holding the capillary array sheet 1 such that the light having passed through the grid pattern enters the through-hole filled with the gel, and an imaging means 16 for imaging images of the grid pattern connected by the gel filled in a plurality of the through-holes of the capillary array sheet. It is characterized by that the imaging means is equipped with an optical system 18 having an angular aperture preventing overlapping of the images of the grid pattern by adjacent through-holes of the capillary array sheet. <P>COPYRIGHT: (C)2010,JPO&INPIT

Description

  The present invention relates to a capillary array sheet inspection apparatus and method, and more specifically, a capillary array sheet inspection apparatus used for detecting biopolymers such as nucleic acids, proteins, polypeptides, polysaccharides, and the like. It relates to the inspection method.

  A capillary array sheet is a kind of biochip (DNA chip or the like) on which a biopolymer probe such as a nucleic acid is immobilized. This capillary array sheet has a plurality of through holes, and a gel in which a biopolymer probe such as a nucleic acid is immobilized is filled in these through holes.

  In the capillary array sheet having such a configuration, a biopolymer probe such as a nucleic acid and a gel precursor solution are injected into a large number of holes formed by a laser beam or the like at the time of manufacture. Due to the entrainment, the gel precursor solution or the like is not completely filled in the holes, and there is a possibility that a through hole in which the gel is insufficient or missing is generated.

  As a capillary array sheet inspection device for discovering through-holes that lack or lack such a gel, the gel filled in the through-holes utilizes the fact that the gel filled in the through-holes acts as a lens. 2. Description of the Related Art A capillary array sheet inspection apparatus and method for determining whether or not a gel is properly filled based on the state of an image that is connected to each other are known (see Patent Document 1).

JP 2003-185597 A

However, the method disclosed in Patent Document 1 irradiates a capillary array sheet with light that has passed through a pinhole, reads a pinhole image connected by a gel filled in a through hole, and reads the read pinhole image. Based on the difference in the state, it is detected whether or not the gel is properly filled (that is, a gel defect), but it takes time to process the image to find the difference in the state of the read pinhole image, It is not suitable for 100% inspection that requires high speed.
In addition, it is difficult to determine the determination criteria for non-defective products and defective products due to the difference in the state of the read pinhole image.

  An object of the present invention is to provide a capillary array sheet inspection apparatus and inspection method capable of reliably and rapidly detecting a gel defect in a through-hole of a capillary array sheet.

According to the present invention,
A capillary array sheet inspection apparatus for inspecting a gel defect of a capillary array sheet having a plurality of through holes filled with gel,
A lighting device with a light source;
A lattice pattern irradiated with illumination light from the light source;
Capillary array sheet holding means for holding a capillary array sheet so that light that has passed through the lattice pattern is incident on a through-hole filled with the gel;
An image pickup means for picking up an image of the lattice pattern connected by the gel filled in the plurality of through holes of the capillary array sheet,
The imaging means includes an optical system having an opening angle at which the images of the lattice pattern formed by adjacent through holes of the capillary array sheet do not overlap.
A capillary array sheet inspection apparatus is provided.

  According to such a configuration, since the optical system of the imaging means has an opening angle such that the lattice pattern images generated by the adjacent through holes do not overlap, an independent lattice pattern image can be obtained for each through hole. Can do. As a result, it is possible to associate a specific through hole with a lattice pattern image connected by the gel of the through hole, and the through hole of the capillary array sheet is filled based on the image obtained by the imaging means. The state of the gel can be determined for each through hole. Therefore, a reliable inspection of the capillary array sheet can be completed in a short time.

According to another preferred embodiment of the present invention, the aperture angle of the optical system is 0.1 to 0.08.
According to another preferred embodiment of the present invention, the aperture angle of the optical system is 0.08.
According to another preferred aspect of the present invention, the grating pattern is an MTF measurement grating.

  According to another preferred aspect of the present invention, when the capillary array sheet holding means can accommodate a liquid therein, the capillary array sheet is held in the capillary array sheet holding means. The liquid is immersed in the liquid in a direction perpendicular to an axis connecting the illumination device and the imaging device.

  According to another aspect of the present invention, each of the capillary array sheets to be inspected is calculated based on the MTF calculated based on the image of the MTF measurement grid imaged by the imaging device of the capillary array sheet inspection apparatus. There is provided a capillary array sheet inspection method characterized by comprising a step of detecting a gel defect in a through hole.

  Hereinafter, a capillary array sheet inspection apparatus and inspection method according to a preferred embodiment of the present invention will be described in detail with reference to the drawings. First, a capillary array sheet in which the absence of a gel is inspected by a capillary array sheet inspection apparatus and inspection method according to a preferred embodiment of the present invention will be described.

  FIG. 1 is a schematic plan view showing the configuration of the capillary array sheet 1. The capillary array sheet 1 includes a plurality of through holes 4 penetrating the sheet-like base material portion 2 in the thickness direction, and a gel 6 on which a biopolymer probe such as a nucleic acid is immobilized is disposed in each through hole 4. Is filled.

  The capillary array sheet 1 is formed by forming a plurality of holes with a laser beam or the like in a plurality of stacked sheet-like members or block-like members, for example, and biopolymer probes such as different nucleic acids and gel precursors in these holes. It is manufactured by injecting a solution, polymerizing the gel precursor solution, and then forming a sheet.

  The capillary array sheet inspection apparatus and inspection method of this embodiment are not limited to the capillary array sheet manufactured by the above-described method, but other methods, for example, biopolymers such as different nucleic acids are immobilized. Capillary manufactured by slicing a fiber assembly (three-dimensional array) formed by regularly arraying a large number of biopolymer-immobilized gel-holding hollow fibers formed in the direction perpendicular to the fiber axis It can also be applied to array sheets.

  FIG. 2 is a schematic drawing showing the configuration of the capillary array sheet inspection apparatus 10 according to a preferred embodiment of the present invention. As shown in FIG. 2, the capillary array sheet inspection apparatus 10 receives an illumination device 14 having a light source 12 such as a halogen lamp and an LED element capable of adjusting the amount of light, and illumination light from the illumination device. The imaging device 16 is provided. The imaging device 16 includes an objective lens 18 that is directed to the illumination device 14 and a CCD sensor 20 that is imaging means. An image obtained by the CCD sensor 20 is sent to an image processing device (not shown) and is subjected to processing such as predetermined image analysis.

  In the capillary array sheet inspection apparatus 10, an MTF (Modulation Transfer Function) measurement grid 22 and an immersion apparatus 24 are arranged between the illumination device 14 and the imaging device 16 in order from the illumination device 14 side. Yes.

  FIG. 3 is a schematic plan view showing an example of the MTF measurement grating 22 used in the capillary array sheet inspection apparatus 1 of the present embodiment. The MTF measurement grating 22 is a well-known MTF measurement grating, and a portion with low light transmittance (black portion) shown in black and a portion with high light transmittance (white portion) shown in white. Are lattice members arranged alternately. In the MTF measurement grid 22, the width of each white portion and each black portion is the same, the width of a pair of white portions and black portions of the lattice pattern is the line pitch (mm), and the reciprocal of the line pitch is the spatial frequency ( lp / mm).

  The lattice pattern of the MTF measurement lattice 22 is formed by a known method such as vapor deposition of a metal on a substrate such as a glass plate. The spatial frequency of the lattice pattern depends on various conditions such as the size of the through hole of the capillary array sheet to be inspected, the thickness of the capillary array sheet to be inspected, and the magnification of the objective lens 18 of the imaging device 16. An appropriate value is selected according to the above.

  The dipping device 24 is a water tank-like member made of a light transmissive material such as glass or quartz, accommodates a liquid 26 such as water, and holds the capillary array sheet 1 in a state dipped in the liquid 26. It is configured to function as a holding device. The surface of the capillary array sheet 1 is oriented in a direction perpendicular to the optical axis of the objective lens 18 of the imaging device 16 (that is, a direction perpendicular to the line connecting the illumination device and the imaging device). It is immersed in the liquid 26 and held in the immersion device 24.

The reason why the capillary array sheet 1 is immersed in the liquid 26 is to prevent the gel 6 held in the through holes 4 of the capillary array sheet 1 from being dried. The liquid may be any liquid as long as it prevents the gel from drying and does not contaminate or deteriorate the gel, and a liquid having high transparency is desirable.
Examples of liquids other than water include salt solutions and glycerol solutions. Moreover, you may add antiseptic | preservative etc. to these solutions.
Further, it is not necessary to provide the immersion device 34 as long as the measurement conditions are such that the gel does not dry. The capillary array sheet 1 only needs to be held in a direction in which the surface thereof is orthogonal to the optical axis of the objective lens 18 of the imaging device 16.

  With such a configuration, the image of the MTF measurement grating 22 illuminated with the illumination light from the illumination device 14 is connected to the image formation position by the lens action of the gel 6 in each through-hole 4 of the capillary array sheet 1. The image is picked up by the CCD sensor 20 of the image pickup device 16 arranged at the image forming position.

  In the present embodiment, the opening angle of the objective lens 18 of the imaging device 16 is such that the image of the MTF measurement grating 22 by the adjacent through holes 4 of the capillary array sheet 1 does not overlap on the CCD sensor 20 of the imaging device 16. That is, the CCD sensor 20 is set so that an independent lattice pattern image for each through hole can be obtained.

  In the present embodiment, the MTF measurement grid 22 is arranged 20 mm away from the illumination device 14, and the capillary array sheet 1 to be inspected is arranged 120 mm away from the MTF measurement grid 22 to obtain the capillary array sheet 1. The image of the MTF measurement grating 22 by the through-holes 4 is set so as to be connected to the imaging position on the rear (imaging means) side of 2 mm from the capillary array sheet 1 so that the imaging position is focused. An objective lens 18 having a focal length of 18 mm, a magnification of 2 times, and an aperture angle (NA) of 0.08 is disposed at a position 18 mm rearward from the imaging position.

  According to such a configuration, the objective lens 18 of the imaging device 16 has a small opening angle such that the MTF measurement grating 22 generated by the adjacent through holes 4 of the capillary array sheet 1 does not overlap. An independent image of the MTF measurement grating 22 can be obtained for each through-hole 4. As a result, it becomes possible to associate the image of the MTF measurement grating 22 connected by the specific through hole 4 and the gel of the through hole 4, and based on the image obtained by the imaging device 16, the capillary array sheet The state of the gel filled in one through hole 4 can be determined for each through hole. Therefore, a reliable inspection of the capillary array sheet 1 can be completed in a short time.

Next, a capillary array sheet inspection method using the capillary array sheet inspection apparatus 10 will be described.
First, the capillary array sheet 1 to be inspected is held at a predetermined position of the immersion apparatus 24 so as to be immersed in the liquid 26 accommodated in the immersion apparatus 24. Next, the illumination light from the illumination device 14 is irradiated onto the MTF measurement grating 22 and the capillary array sheet 1, and an image of the MTF measurement grating 22 is generated by the imaging device 16.

  The image of the MTF measurement grating corresponding to each through hole obtained by the CCD sensor 20 is processed by the processing device, and the MTF for each through hole is calculated from the light amount line profile for each through hole according to the following equation 1. The determination criteria (threshold values) of the MTF and the gel filling state are compared, and the suitability of the gel filling state is determined for each through hole.

MTF = {(Imax-Imin) / (Imax + Imin)} × 100 (%) Equation 1

  Here, Imax and Imin are values for calculating the MTF, and are the maximum light amount value (Imax) and the minimum light amount value (Imin) of the light amount line profile of the image of the imaged MTF measurement grating. .

  As described above, in this embodiment, since the numerical aperture of the objective lens 18 is small, as shown in FIG. 4, the lattice image incident on the objective lens 18 and formed by one through-hole 4 at the imaging position is shown. The range is narrowly limited as indicated by the solid line compared to the range incident on the objective lens having a large aperture angle indicated by the dotted line. As a result, an independent image of the MTF measurement grating 22 can be obtained for each through-hole 4.

Without being limited to the above-described embodiment of the present invention, various changes and modifications can be made within the scope of the technical idea described in the claims.
In the above embodiment, the aperture angle of the objective lens is 0.08, but the imaging means is limited to the above numerical value as long as the aperture angle does not overlap the image of the lattice pattern by the adjacent through holes of the capillary array sheet. It can be changed depending on the magnification of the lens, the size of each member, the mutual distance, etc., and may be in the range of 0.01 to 0.08, for example.

  Examples of the present invention will be described below.

Capillary array sheet fabrication plate Using two perforated plates with a thickness of 0.1 mm in which 19 × 12 and 228 holes having a diameter of 0.32 mm are arranged at intervals of 0.42 mm and arranged in a lattice form at the center of the plate. A porous plate made of polycarbonate colored with carbon black (outer diameter 0.28 mm, inner diameter 0.18 mm, length 500 mm) is passed through all the holes of the perforated plate, and the two plates are passed through the perforated plate. 50 mm apart, filled with a polyurethane resin colored with carbon black between the separated perforated plates, and a hollow fiber array having a prismatic shape with a length of 50 mm and a square of 7 × 12 mm, having portions not fixed by the resin at both ends Got.

  The hollow fiber in the obtained hollow fiber array is filled with the gel precursor solution and 40-type nucleic acid probes of different types while entraining bubbles, polymerizing the gel precursor solution, and then with a microtome, A capillary array sheet was obtained by cutting out a 0.25 mm-thick slice in a direction perpendicular to the fiber axis.

(Inspection equipment)
The obtained capillary array sheet was immersed in a glass petri dish containing sterilized water and set on the stage of a microscope (Olympus BX60). For the microscope, the condenser was removed, the objective lens used was a magnification of 2.times., The aperture angle (NA) was 0.08, and the illumination device was transmitted illumination (halogen lamp) attached to the microscope.
As shown schematically in Fig. 5, an MTF measurement grid (2 lp / mm, thickness 1 mm) manufactured by depositing metal on a glass plate is attached 120 mm below the capillary array sheet. A light source was placed 20 mm below the grid.

(Inspection)
When the focus handle was manually operated, a lattice image was formed 2mm above the capillary array sheet surface. An image of the lattice image of the MTF measurement lattice formed through each through-hole of the capillary array sheet was captured by a CCD sensor (COOLSNAP manufactured by Photometrics). This image is shown in FIG. 6 and FIG. 7 in which a part of FIG. 6 is enlarged. According to the present embodiment, as shown in FIGS. 6 and 7, a lattice image for each through hole 4 of the capillary array sheet 1 can be obtained independently. As a result, the gel filling state can be easily and reliably determined for each through-hole based on the lattice image.

(Comparative example)
FIG. 9 is an enlarged view of a part of FIG. 8 and FIG. 8 showing an image captured under the same conditions as in the above example except that an objective lens having a magnification of 4 times and an aperture angle (NA) of 0.13 is used. Show. As shown in FIGS. 8 and 9, the lattice image by the through holes 4 of the capillary array sheet 1 overlaps with the lattice image by the adjacent through holes in the peripheral portion.

It is a typical top view which shows the structure of an example of a capillary array sheet | seat. 1 is a schematic drawing showing a configuration of a capillary array sheet inspection apparatus according to a preferred embodiment of the present invention. FIG. 3 is a schematic plan view showing an example of an MTF measurement grating used in the capillary array sheet inspection apparatus of FIG. 2. 3 is a diagram illustrating a range of an image according to an embodiment of the present invention. It is a figure which shows typically the inspection method of the Example of this invention. It is a photograph of the image obtained in the Example of this invention. FIG. 7 is an enlarged view of a part of FIG. 6. It is a photograph of the image obtained in the comparative example of this invention. FIG. 9 is an enlarged view of a part of FIG. 8.

Explanation of symbols

1: Capillary array sheet 2: Base material part 4: Through hole 6: Gel 10: Capillary array sheet inspection device 14: Illumination device 16: Imaging device 20: CCD sensor 22: MTF (modulation transfer function) measurement Grid 24: Immersion device

Claims (6)

A capillary array sheet inspection apparatus for inspecting a gel defect of a capillary array sheet having a plurality of through holes filled with gel,
A lighting device with a light source;
A lattice pattern irradiated with illumination light from the light source;
Capillary array sheet holding means for holding a capillary array sheet so that light that has passed through the lattice pattern is incident on a through-hole filled with the gel;
An image pickup means for picking up an image of the lattice pattern connected by the gel filled in the plurality of through holes of the capillary array sheet,
The imaging means includes an optical system having an opening angle at which the images of the lattice pattern formed by adjacent through holes of the capillary array sheet do not overlap.
Capillary array sheet inspection apparatus. An aperture angle of the optical system is 0.1 to 0.08;
The capillary array sheet inspection apparatus according to claim 1. The aperture angle of the optical system is 0.08.
The capillary array sheet inspection apparatus according to claim 2. The lattice pattern is an MTF measurement lattice.
The capillary array sheet inspection apparatus according to any one of claims 1 to 3. The capillary array sheet holding means is capable of containing a liquid therein, and when the capillary array sheet is held in the capillary array sheet holding means, a line connecting the illumination device and the imaging device is formed. Immersed in the liquid in an orthogonal orientation;
The capillary array sheet inspection apparatus according to any one of claims 1 to 4. A capillary array to be inspected is calculated based on an MTF calculated based on an image of an MTF measurement grid imaged by the imaging device of the capillary array sheet inspection apparatus according to any one of claims 1 to 5. Detecting a gel defect in each through-hole of the sheet,
Capillary array sheet inspection method characterized by the above.

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