KR100717819B1 - Spotting device for microarraying biological material - Google Patents

Abstract

The present invention relates to a spotting device for the microarray of a biological sample. More specifically, the spotting device for the micro-arrangement of biological samples comprising a plurality of pins and pin guiding holder to contact and support the pins at a plurality of points to allow the pins to perform a precise linear movement in the vertical direction will be.

Pin holder

Description

Spotting device for microarraying biological material

1 is an exploded perspective view of the present invention.

2 is a schematic view of the pin of the present invention.

<Short description of the symbols in the drawings>

1: upper guide plate 2: lower guide plate

3: guide wire 4: adjusting block

5: frame 6: spring plate

7: adjusting screw 8: pin

The present invention relates to a spotting device for the microarray of a biological sample. More specifically, the spotting device for the micro-arrangement of biological samples comprising a plurality of pins and pin guiding holder to contact and support the pins at a plurality of points to allow the pins to perform a precise linear movement in the vertical direction will be.

As the human genome project is completed, a large amount of genomic information is continuously revealed, and based on the nucleotide sequence information of the genes identified, development of a technique for efficiently interpreting the functions of genes of various organisms is required. Accordingly, functional genomics or system biology has emerged as a post-genome research project, and these are studies for commercial application of the results of the human genome project. An example of such studies is the study of DNA chips in which a large number of DNA molecules are arranged on a substrate such as slide glass or silicon.

So-called DNA chip technology, also known as DNA microarray technology, is a very useful technique for simultaneous interpretation of specific gene expression, mutations, and polymorphisms. Very suitable technique (Lipshutz, RJ et al. (1995) Biotechniques 19, 442-447; Chee, M. et al. (1996) Science 274, 610-614).

DNA chips are manufactured by existing molecular biological knowledge and mechanical and electronics technology. DNA chips integrate hundreds to hundreds of thousands of DNA into small spaces using automated mechanical devices and electronic control techniques. Using a DNA chip that has accumulated hundreds to hundreds of thousands of DNA types, at least hundreds of genes can be searched in a short time at a time. The DNA chip uses a glass plate or a semiconductor wafer as a substrate, and since a very small amount of hundreds of thousands of DNAs are highly integrated and arranged, many kinds of genes can be searched in a short time at a time.

To date, various methods of manufacturing DNA chips have been developed, and are classified into spotting or contact printing, non-contact printing, and photo-lithography.

Among these methods, the non-contact printing method is a method of quantitatively spraying a DNA solution onto a substrate by applying heat or electrical force to the DNA solution reservoir using a solenoid actuator or piezoelectric element. However, in general, since there are many kinds of DNA solutions to be injected onto the DNA chip, a large number of reservoirs are required, and the remaining space present in the channel serves as the injection pressure buffering function and it is difficult to elaborate precise metering.

Photo-lithography is a technique used in the current semiconductor manufacturing process, a method of synthesizing DNA oligomers on the surface of the substrate using a protecting group removed by ultraviolet (UV), the base required Arrays of DNA oligomers can be formed directly on a substrate (Pease, AC et al. (1994) Proc. Natl. Acad. Sci. USA 91, 5022-5026). However, this method is a technique that cannot be easily used by ordinary researchers because it requires the use of complicated equipment. In addition, while high density integration is possible, the desired DNA length is limited to about 20 base pairs or less.

The spotting or contact printing method is a method in which a fine pin such as a pointed needle is immersed in a DNA oligomer solution, followed by contacting the end of the pin on a slide glass substrate to drop a small amount of DNA oligomer solution. In the case of using a needle-shaped pin without an internal space for storing the DNA oligomer solution, the device needs to be immersed in the DNA oligomer solution every time spotting, so that some special arrayer or colony picking robots are used. Only limited use can be made. In order to overcome the limitation of the use of needle-shaped pins, a quill method using a grooved pin having a width of about 100 microns inside and outside of the pin has been proposed (DeRisi, JL et al. (1997). ) Science 270, 680-686, US Pat. No. 6101946).

The pin of this shape, when the pin is immersed in the DNA oligomer solution, the micro oligomer solution of several microliters is sucked up by the capillary phenomenon, it is possible to repeatedly spot the DNA oligomer solution on a plurality of substrates. However, quill pins are very difficult to process. In addition, a method of spotting sequentially on a solid substrate using a plurality of glass capillaries having an inner diameter of 100 to 200 μm has been reported. To form spots of about 100 μm diameter on the slide glass, fine pins with corresponding diameter ends must be used. In addition, in order to form a high density spot with a diameter of 100 μm in an area of 1 cm 2, precise linear motion in the vertical direction of the spotting pin is required.

In this pin spotting method, a plurality of pins having end portions having an inner diameter of 50 to 300 μm are mounted in a holder, and then the holder is moved vertically and horizontally so that the ends of the pins are immersed in the DNA oligomer solution to move the DNA oligomer solution into the pins. After inhalation, the inhaled DNA oligomer solution is dipped onto the substrate to form a DNA oligomer spot. In such a pin spotting method, since the DNA oligomer solution is deposited on the substrate by the vertical movement of the pin holder, a mechanical device for precisely controlling the vertical operation of the holder is required.

However, the spotting apparatus used in the DNA microarray equipment of the pin spotting method, which is known so far, has a fine straight line in which the pin is vertically moved according to the operation of the holder so that the DNA oligomer solution spot is densely arranged on the substrate. Despite the need to move, the pins are inclined somewhat in the holder, so that the linear movement is not precisely up and down, and as a result, the position where the spot is formed deviates from the intended position or overlaps with another spot. there was. In addition, the conventional pin spotting device has a problem in that the size of the spot formed on the substrate becomes inconsistent or the pin is separated after contact with the substrate, causing a phenomenon that the pin slips out of the target position, making it difficult to manufacture a sophisticated high density fine array. .

Therefore, in the art, a micro-myararrayer using a large number of pins, in order to form fine spots on a narrow substrate with high density, spotting pins can be inserted to allow precise linear movement in the vertical direction. The development of pin spotting devices has long been expected.

Accordingly, the present invention is a device for solving the problems of the conventional pin spotting device, pin guiding holder to secure the position reproducibility of spots deposited by the spotting pin and to enable precise linear movement of the pin up and down direction And to provide a spotting device consisting of a pin suitable for being inserted into it.

Spotting device for biological sample microarray of the present invention

A large number of pins 8 that suck the biological sample and finely drip it onto the substrate;

Upper guide plate (1) with pin insertion holes forming two pin support points,

A plurality of guide wires (3) supporting the inserted pins,

Adjusting block (4) is the guide wire is inserted into a plurality of rows up and down,

A spring plate 6 for pushing the adjusting block 4 inserted into the frame 5 in the direction in which the adjusting screw 7 is installed;

Adjustment screw (7) for pushing the adjustment block 4 in the direction in which the spring plate 6 is installed,

The frame 5 is inserted into the adjustment block, the spring plate 6 is inserted, and the adjustment screw 7 is mounted;

It is characterized in that it comprises a lower guide plate (2) having pin insertion holes forming two pin support points.

The guide wire 3 is made of a material having an elastic or metal material to perform the function of adjusting the strength of the force supporting the pin through the adjustment of the adjusting screw (7).

Hereinafter, embodiments of the spotting apparatus of the present invention of the present invention will be described in more detail with reference to the accompanying drawings. The spotting apparatus of the present invention shown in the accompanying drawings is for explaining one example of the spotting apparatus of the present invention, and the content of the present invention is not limited to the apparatus shown in the following drawings.

1 is an exploded perspective view of a spotting device for the micro array of biological samples according to the present invention. A spring plate 6 is mounted on one of the inner surfaces of the frame 5, and a hole in which the adjusting screw 7 for adjusting the position of the adjusting block is tightened is formed on the inner wall facing the inner surface. Guide plates 1 and 2 are attached to the upper and lower surfaces of the frame 5 of the present invention, and the adjustment block 4 is mounted in the frame 5.

As shown in FIG. 1, the adjusting block 4 has a plurality of guide wires 3 formed in a transverse direction through which the pin 8 is inserted, and having a cylindrical support and one contact point. It is inserted side by side. More specifically, the guide wires 3 are arranged at regular intervals across the adjusting block 4, and are inserted into the adjusting blocks by being inserted into respective holes formed in a plurality of rows in the wall of the adjusting block. Preferably, a hole is formed in two rows of upper and lower rows on the remaining wall where the spring plate 6 or the adjusting screw 7 hole is not formed among the four wall surfaces of the adjusting block, and the guide wire is inserted into each hole. When the pin is inserted in the vertical direction, the pin is in contact with the two upper and lower guide wires, respectively.

Thus, the pin is inserted into the adjusting block and supported in contact with the two guide wires mounted to the adjusting block. The adjusting block is mounted in the frame and fixed by a spring 7 and a screw 7 for adjusting the position of the adjusting block.

The spring plate 6 is installed between the adjusting block and the frame to push out the frame and the adjusting screw installed on the opposite surface adjusts the left and right positions of the adjusting block in the frame. Typically, the size of the control block mounted in the frame is smaller than the size of the frame to adjust the position of the left and right by the spring plate and screws in the frame.                     

In this case, the position of the adjusting block in the frame can be adjusted to adjust the bearing force applied to the pin by the guide wire in contact with the pin.

The guide plate consists of an upper guide plate 1 and a lower guide plate 2 attached to the upper and lower surfaces of the frame, respectively. The pin insertion hole formed in the upper and lower guide plates is formed by a circular or polygonal hole and has two or more contact supporting points with the pins, respectively. In order to form such a contact support point, as shown in FIG. 1, a hole of a triangular to octagonal shape, most preferably a triangular shape is formed. The hole shape formed in the guide plate may be various shapes such as circular or polygonal, and the number of holes may be adjusted according to the number of pins to be inserted.

In one embodiment of the present invention, the guide plate is formed with 32 triangular holes so that 32 pins can be mounted. According to the number of pins used, the number of holes formed in the guide plate is appropriately adjusted. As shown in Figure 1, the hole is an equilateral triangle, a total of 32 horizontally formed, eight horizontally, four vertically. The pin is inserted into a triangular hole formed in the upper and lower guide plates to contact at two points of the corner of the hole.

2 is a schematic view of a pin according to the present invention. The pins come in contact with the edges of the triangular holes of the upper guide plate at two points, with two rows of wires attached to the control block in the frame, and at the two points with the edges of the triangle holes of the lower guide plate. A total of six points are supported. In order to fix the pin to the holder by applying a supporting force at the contact point, the adjusting screw is moved left and right to adjust the position of the adjusting block.                     

Since the pin is in contact with the four fixed support points of the hole formed in the guide plate, it is supported by the guide wire, which is a material having elastic or metallic material, so that the supporting force applied to the pin is controlled by the guide wire.

By operating the pin-mounted holder in the up and down direction, the pin can be precisely moved vertically without tilting in the holder.

As the contact point between the pin and the holder increases and the contact area increases, the friction force between the pin and the contact surface increases, which not only causes the pin to wear easily, but also causes problems in vertical movement of the pin due to the friction force during vertical movement of the pin. Can be. In the spotting apparatus of the present invention, the pin support point in contact with the pin is five (5) or more, preferably six (6), so that the frictional force of the pin can be minimized and the pin is properly fixed. Pin can smoothly elaborate linear movement up and down.

As the pin of the present invention, a capillary pin or a stealth pin may be used. The diameter of the pin is very important because it determines the size of the spot formed in the chip manufacturing. In the present invention, preferably capillary pins may be used, capillary pins having an inner diameter of 0.5 to 3 mm are used, and capillary pins of 0.5 mm to 2 mm are usually used.

In addition, the spotting apparatus of the present invention can be used by attaching not only a capillary pin but also a stealth pin of various forms such as a well-known needle (quill type) having a shape such as a fountain pen nib and a deformation thereof.

In the present invention, "stealth pin" is designed by Brown (US Patent Nos. 6,110,426 and 5807522 to Brown) and Martinsky (US Patent No. 6,101,946 to Martinsky). It refers to a pin that is formed and has an opening formed to penetrate the pin while being connected to the cutout of the pin. These steel spins are metal capillary pins made of stainless steel and have a fountain pen nib shape, and commercially available steel spins include Arraylt ^™ pins commercially available from TeleChem International Inc. of the United States.

Preferably, a stopper is inserted at the opposite end of the tip portion of the pin to install each spotting pin into the holder. The stopper may be inserted at the end of the pin, or may be inserted at a portion of the pin. In the present invention, the material of the stopper is made of rubber of elastic material to make a hole in the center, and a pin is inserted therein. As the material of the stopper, other elastic materials, metals, alloys, or the like can be used.

In the present invention, although the holder is mounted with eight 32 horizontally, four pins vertically, but is not limited to this, the holder is mounted 16 vertically eight 128 pins can also be used. In the present invention, when using 32 pins, a 384 well plate is appropriately used to finely arrange a sample in a short time. When using 128 pins, a 1536 well plate is used to finely arrange a larger amount of a sample in a short time. can do. When the spotting pin is immersed in the well containing the biological sample, the sample is sucked into the pin, and the pin containing the sample is moved onto the substrate by the xyz Cartesian robot arm and brought into contact with the sample to form a spot on the substrate.

As described above, the spotting apparatus of the present invention includes a plurality of pins and a spot guiding device for micro array of biological samples including a pin guiding holder for contact supporting each of the plurality of pins at five or more places. As it minimizes the contact between the pin and the holder, the friction between the pin and the holder is minimized, which improves the durability of the pin, secures the positional reproducibility of the pin and enables the precise linear movement of the pin in the vertical direction. It provides the effect of enabling a fine microarray of the sample.

Claims (9)

A large number of pins 8 that suck the biological sample and finely drip it onto the substrate; Upper guide plate (1) with pin insertion holes forming two pin support points, A plurality of guide wires (3) supporting the inserted pins, Adjusting block (4) is the guide wire is inserted into a plurality of rows up and down, A spring plate 6 for pushing the adjusting block 4 inserted into the frame 5 in the direction in which the adjusting screw 7 is installed; Adjustment screw (7) for pushing the adjustment block 4 in the direction in which the spring plate 6 is installed, The frame 5 is inserted into the adjustment block, the spring plate 6 is inserted, and the adjustment screw 7 is mounted; Spotting device for micro array of biological samples, characterized in that it comprises a lower guide plate (2) having pin insertion holes forming two pin support points. The spotting apparatus of claim 1, wherein the guide wire is made of an elastic or metallic material. The spotting device of claim 1, wherein the pins are capillary pins. The spotting apparatus of claim 1, wherein the guide wire is inserted into holes formed in two rows of upper and lower rows of the control block. The spotting apparatus of claim 1, wherein the pin is inserted into and mounted on the stopper. The spotting apparatus of claim 5, wherein the stopper is made of rubber of an elastic material. The spotting apparatus of claim 1, wherein the pin is a steel spin. 8. The spotting device of claim 7, wherein the pin is inserted into and mounted on the stopper. The spotting apparatus of claim 8, wherein the stopper is made of rubber of an elastic material.

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