CN218710327U - Space transcriptome biochip - Google Patents

Abstract

The utility model relates to the field of biotechnology, a space transcriptome biochip is provided, include: the transparent substrate forms a micropore area through photoetching and etching, and a micropore structure is formed in the micropore area and used for placing the coding microspheres with the primers. The utility model provides a space transcriptome biochip, because of adopting the transparent basement, it can carry out the light field microscopic imaging of HE dyeing, toluidine blue dyeing and masson's pine dyeing etc., combine gene expression result and light field dyeing result to carry out contrastive analysis; in addition, the micro-pore structure of the chip has high resolution, and can realize the analysis of the space transcriptome data at the sub-cell level.

Description

Space transcriptome biochip

Technical Field

The utility model relates to the field of biotechnology, especially, relate to a space transcriptome biochip.

Background

The pattern of gene expression at the original location of the space in the tissue is important for understanding the type and function of the cells therein. In recent years, space transcriptome technology has been rapidly developed and widely applied to various fields such as tumors, diseases, development of nervous systems and organs, and the like.

The carriers currently used for biochip fabrication generally have reactive groups that can undergo chemical reactions for coupling biomolecules. Types of such carriers mainly include glass slides, silicon wafers, nitrocellulose membranes, nylon membranes, and the like. The following two types of spatial transcriptome chips are available in the prior art:

1. the glass slide is used, and the durable carrier can resist high temperature and high ionic strength and has non-wettability, so that the hybridization volume is reduced to the minimum; in addition, the hydrophobic surface overcomes the defect that the sample is easy to diffuse, and the density of the sample points is improved; the low fluorescence signal does not cause strong background interference. The biochip is used for fixing capture probes for spatial analysis on a slide, but has the defect of low resolution of sample analysis, and cannot meet the requirement of scientists on subcellular structure analysis.

2. The chip made of the material is developed based on gene sequencing, and can realize the analysis of the space transcriptome at the level of subcellular, but the substrate made of non-transparent material can not carry out bright field microscopic imaging such as HE staining, toluidine blue staining, masson staining and the like, and can only realize the imaging of tissues in a fluorescence mode or use clinical pictures to carry out the bright field imaging, so that the final gene expression result can not be well combined with the microscope acquisition result for analysis.

SUMMERY OF THE UTILITY MODEL

The utility model provides a space transcriptome biochip for solve among the prior art biochip adoption non-transparent material's basement can't carry out HE dyeing, toluidine blue dyeing and the micro-defect of formation of image of bright field such as masson's dyeing and adopt the slide basement can't satisfy the defect of scientist to subcellular structure analysis demand.

The utility model provides a space transcriptome biochip, include:

the transparent substrate forms a micropore area through photoetching and etching, and a micropore structure is formed in the micropore area and used for placing the coding microspheres with the primers; the micropore area forms at least one sub-area, and the sub-area is a circular area or a polygonal area.

According to the utility model provides a pair of space transcriptome biochip, form a plurality of small units in the subregion, it is adjacent the interval sets up between the small unit, small unit is according to circular or polygon repetition arrangement.

According to the utility model provides a pair of space transcriptome biochip, the edge and/or the angle of small unit form the breach to form the position mark.

According to the utility model provides a pair of space transcriptome biochip, the distance is between 0 micron-40 micron between the small unit.

According to the utility model provides a pair of space transcriptome biochip, microporous structure's diameter value range is between 0.1 micron to 10 microns, and two adjacent micropore centre points are apart from between 0.1 micron to 20 microns.

According to the utility model provides a pair of space transcriptome biochip, the long limit value scope of transparent basement is 10-100mm, and the minor face value scope is 5-50mm.

According to the utility model provides a space transcriptome biochip, the size value range of subregion is between 9mm x 9mm to 1875mm.

According to the utility model provides a pair of space transcriptome biochip, transparent basement is one of glass, quartz, plastics, magnesium chloride and gallium arsenide.

According to the utility model provides a pair of space transcriptome biochip, the microporous structure is including being located transparent basement is on the surface flaring portion and being located the inside throat portion of transparent basement, the flaring portion with the throat portion along the depth direction intercommunication of transparent basement is in order to form on the transparent basement microporous structure.

According to the utility model provides a space transcriptome biochip, the transparent substrate has a first surface and a second surface, the first surface and the second surface form a micropore area through photoetching and etching respectively;

wherein the first surface and the second surface are two opposite surfaces of the transparent substrate.

The utility model provides a space transcriptome biochip, it has adopted transparent substrate, utilize photoetching and etching technique to make the cellular structure and form the micropore area, its preparation simple process, reduce the material consumption cost, and can image through the bright field, combine with gene expression result, greatly improved the analytic effect of space transcriptome; in addition, the micro-pore structure of the chip has high resolution, and can realize the analysis of the space transcriptome data at the sub-cell level.

Drawings

In order to more clearly illustrate the technical solutions of the present invention or the prior art, the following briefly introduces the drawings required for the embodiments or the prior art descriptions, and obviously, the drawings in the following description are some embodiments of the present invention, and for those skilled in the art, other drawings can be obtained according to the drawings without creative efforts.

FIG. 1 is a schematic structural diagram of a space transcriptome biochip having a neutron region in the form of a large rectangle;

FIG. 2 is a schematic structural diagram of a space transcriptome biochip having a small rectangular neutron region;

FIG. 3 is a schematic diagram of a spatial transcriptome biochip having a circular neutron region;

FIG. 4 is a schematic diagram of a structure in which the neutron region of the space transcriptome biochip according to the present invention is hexagonal;

FIG. 5 is a schematic diagram of the structure of the spatial transcriptome biochip with rectangular and uniform micro-units;

FIG. 6 is a schematic structural view of the spatial transcriptome biochip according to the present invention in which the micro-units are arranged in a hexagonal pattern;

FIG. 7 is a schematic structural diagram of a spatial transcriptome biochip according to the present invention, wherein the gap is located at the edge of the micro-unit;

FIG. 8 is a schematic structural diagram of the position of the gap in the micro-unit of the space transcriptome biochip according to the present invention;

FIG. 9 is a schematic structural view of the space transcriptome biochip according to the present invention without gaps formed in the micro-cells;

FIG. 10 is a longitudinal sectional view of a space transcriptome chip provided by the present invention.

Reference numerals are as follows:

10: a transparent substrate; 11: a first surface; 12: a second surface; 20: a microporous region; 30: a sub-region; 40: a minute unit; 50: a notch; 60: a gap; 70: a microporous structure; 71: a flared part; 72: a necking-down portion.

Detailed Description

To make the objects, technical solutions and advantages of the present invention clearer, the drawings in the present invention will be combined to clearly and completely describe the technical solutions of the present invention, and obviously, the described embodiments are some, but not all embodiments of the present invention. Based on the embodiments in the present invention, all other embodiments obtained by a person skilled in the art without creative efforts all belong to the protection scope of the present invention.

A spatial transcriptome biochip of the present invention is described below with reference to FIGS. 1 to 9, comprising: a transparent substrate 10.

The transparent substrate 10 is formed with a micropore area 20 by photolithography and etching, a micropore structure 70 is formed in the micropore area 20, and the micropore structure 70 is used for placing the encoded microspheres with primers.

Further, the transparent substrate 10 in the present embodiment may employ glass, quartz, plastic, or transparent conductive plating; the transparent conductive coating can be one of magnesium chloride, gallium arsenide and the like. The micropore area 20 is formed on the transparent substrate 10 by means of photolithography and etching, and the micropore structure 70 in the micropore area 20 can be used for placing the encoded microspheres with primers.

The microporous region 20 in this embodiment may be a microporous region 20 formed by uniformly tiling the microporous structure 70 as a whole, or may also be formed into sub-regions 30 of different shapes (specifically illustrated in the following embodiments), and the sub-regions 30 of different shapes may be more beneficial to different scanning software to perform image stitching and identification, and the like. The resolution of the spatial transcription component is improved through the microporous structure 70, high-sensitivity detection is realized, and the requirement of scientists on subcellular structure analysis is met; moreover, the glass substrate can be adopted, the micropore structure 70 can be manufactured by utilizing the etching technology, the manufacturing process is simple, and the material consumption cost is reduced.

The space transcriptome biochip in the embodiment can be used for performing bright field microscopic imaging of HE staining, toluidine blue staining, masson staining and the like, and can be used for performing comparative analysis by combining a gene expression result and a bright field staining result.

The utility model provides a space transcriptome biochip, it has adopted transparent basement, utilizes photoetching and etching technique to make the micropore structure and form the micropore area, its preparation simple process, reduces the consumptive material cost, and can pass through the bright field formation of image, combine with gene expression result, greatly improved the analysis effect of space transcriptome; in addition, the micropore structure of the chip has high resolution, and can realize the analysis of the space transcriptome data at the subcellular level.

In one embodiment of the present invention, the microporous region 20 forms at least one subregion 30, and the subregion 30 is a circular region or a polygonal region. It is understood that the sub-region 30 in this embodiment is also formed by a plurality of micro-porous structures 70, and the plurality of sub-regions 30 form a whole micro-porous region 20 according to a specific arrangement. For example, the sub-regions 30 are shown in FIG. 1 as one large rectangle, in FIG. 2 as multiple rows and columns of small rectangles, in FIG. 3 as two circular arrays of sub-regions 30, and in FIG. 4 as four hexagons of sub-regions 30. It is understood that the sub-region 30 may be one as shown in fig. 1, or may be a plurality of the sub-regions as shown in fig. 2, 3 and 4, and the shape of the sub-region 30 may be not only a circle, but also a polygon such as a triangle, a rectangle, a hexagon, etc., which is not exemplified herein.

In one embodiment of the present invention, a plurality of small units 40 are formed in the sub-region 30, and the adjacent small units 40 are spaced apart from each other, and the small units 40 are uniformly arranged or repeatedly arranged in a circular or polygonal shape. In the present embodiment, a plurality of smaller cells 40 are also formed in the sub-region 30, and the cells 40 in the same sub-region 30 are uniformly arranged in a circle or a polygon, for example, the cells 40 may be uniformly arranged in a rectangle as shown in fig. 5, or may be distributed in a hexagon as shown in fig. 6. It is understood that the minute unit 40 is also constituted by a plurality of the minute structures 70, and the plurality of minute units 40 constitute one of the minute regions 30 in a specific arrangement. In the prior art, when a scanning device scans the microporous structure 70 on the substrate, the scanned images need to be spliced by using a software algorithm, and the scanned images at the previous and subsequent times need to be spliced in the splicing process, but due to the numerous microporous structures 70, the problem of splicing errors often occurs in the splicing process. In this embodiment, a plurality of gaps 60 are arranged in the sub-region 30, so that the sub-region 30 is divided into a plurality of small-sized micro units 40, the images in different regions can be conveniently identified by scanning equipment and software, the correction of image acquisition in the later period is facilitated, the positioning error of chip coding information is avoided, and the splicing accuracy is improved.

Further, if the sub-region 30 is square, the size of the sub-region 30 ranges from 9mm × 9mm to 1875mm × 1875mm, and of course, the size of the sub-region 30 may be adjusted according to different shapes and different experimental requirements. The distance between the minute cells 40 is 0 to 40 micrometers, and the minute cells 40 are similarly,

in one embodiment of the present invention, the edges and/or corners of the tiny cells 40 form indentations 50 to form orientation marks. Therefore, in the embodiment, the orientation mark is arranged at the edge and/or the angular position of the tiny unit 40 located at the edge and/or the angular position of the sub-region 30, so that the orientation is conveniently distinguished, the rotation direction and the angle of the image are identified, and the accuracy of image processing is further improved. Specifically, the minute cell 40 provided with the notch 50 in the present embodiment refers to the minute cell 40 located at the edge and/or the angular position of the sub-area 30, the shape of the notch 50 may be rectangular, triangular, etc., and the notch 50 is a part cut out of the minute cell 40 to form an orientation mark for distinguishing the orientation and identifying the rotation direction and angle of the image. Since the shape of the tiny unit 40 can be circular or polygonal, the gap 50 can be located at the edge of the tiny unit 40 as shown in fig. 7; or the notch 50 may be located at the corner of the tiny cell 40 as shown in fig. 8; fig. 9 is a schematic view showing that the notch 50 is not formed in the minute cell 40.

In one embodiment of the present invention, the diameter of the microporous structure 70 ranges from 0.1 micrometer to 10 micrometers, and the distance between the center points of two adjacent micropores ranges from 0.1 micrometer to 20 micrometers. In the present embodiment, the microporous structure 70 is formed inside the microporous region 20 formed by tiling the microporous structures 70, or the microporous region 20 formed by the sub-regions 30 and the micro units 40, the microporous structure 70 may be in the form of circular micropores, the diameter of the microporous structure is between 0.1 micron and 10 microns, the center point distance between two adjacent micropores is between 0.1 micron and 20 microns, and of course, according to different requirements, the size of the diameter of the micropores and the size of the center point distance of the micropores may also be corresponded.

In one embodiment of the present invention, the long side of the transparent substrate 10 ranges from 10 to 100mm, and the short side ranges from 5 to 50mm. For example, the length, width, and height of the transparent substrate 10 may be 75mm 25mm 1mm, and the size of the transparent substrate 10 may be designed accordingly according to actual needs.

In one embodiment of the present invention, the microporous structure 70 includes a flared portion 71 located on the surface of the transparent substrate 10 and a necked portion 72 located inside the transparent substrate 10, and the flared portion 71 and the necked portion 72 communicate along the depth direction of the transparent substrate 10 to form the microporous structure 70 on the transparent substrate 10. In the present embodiment, the microporous structure 70 has a deep taper structure in a funnel shape in the depth direction, and a diameter of a flared portion 71 formed on the surface of the transparent substrate 10 is larger than a diameter of a necked portion 72 formed inside the transparent substrate 10, that is, the diameter of the microporous structure 70 is gradually reduced from the surface of the transparent substrate 10 to the inside. By processing the microporous structure 70, the diameter of the flared part 71 is larger, so that the encoded microspheres are positioned conveniently and are easier to place. Further, the oblique angle of the deep cone structure is between 0 ° and 60 °, that is, the included angle between the connecting line of the flared portion 71 and the necking portion 72 and the depth direction of the transparent substrate 10 is between 0 ° and 60 °, and of course, the angle can be adjusted according to actual needs.

In one embodiment of the present invention, the transparent substrate 10 has a first surface 11 and a second surface 12, and the first surface 11 and the second surface 12 are respectively formed with the micro-pore region 70 by photolithography and etching; the first surface 11 and the second surface 12 are two opposite surfaces of the transparent substrate 10. In this embodiment, by processing the micro-porous regions on both the front and back surfaces (i.e., the first surface 11 and the second surface 12) of the transparent substrate 10, when the encoded microspheres are placed on the first surface 11 (i.e., the front surface), the second surface 12 (i.e., the bottom surface) can be subjected to other processes, such as pre-treatment or post-treatment, such as cleaning, of the micro-porous regions, thereby improving the overall working efficiency.

The utility model provides a size design of space transcriptome biochip, the length of glass basement, width and thickness are 75mm, 25mm and 1mm respectively, arrange eight sub-regions of rectangle 30 on the glass basement, and set up four rows along length direction, set up two along width direction, sub-region of rectangle 30 is 8mm with the distance at glass basement top, be 3.3mm with the distance of the side of glass basement, two are 4mm apart between the sub-region of rectangle 30, sub-region of rectangle 30 before the adjacent row is 5mm apart. The length and width of each rectangular sub-region 30 are 7.2mm and 7.2mm respectively. The aperture of the micro-pore structure 70 etched on the transparent substrate 10 is about 2.5um, which is equivalent to that the resolution of the current mainstream space transcription component is improved by more than 20 times, and HE staining and gene expression experiments can be performed on a glass slide at the later stage.

Finally, it should be noted that: the above embodiments are only used to illustrate the technical solution of the present invention, and not to limit it; although the present invention has been described in detail with reference to the foregoing embodiments, it should be understood by those skilled in the art that: the technical solutions described in the foregoing embodiments may still be modified, or some technical features may be equivalently replaced; such modifications and substitutions do not depart from the spirit and scope of the present invention in its corresponding aspects.

Claims (10)

1. A spatial transcriptome biochip, comprising:

the transparent substrate forms a micropore area through photoetching and etching, and a micropore structure is formed in the micropore area and used for placing the coding microspheres with the primers; the micropore area forms at least one sub-area, and the sub-area is a circular area or a polygonal area.

2. The spatial transcriptome biochip of claim 1, wherein a plurality of micro units are formed in the sub-region, and are spaced apart from each other, and the micro units are repeatedly arranged in a circular or polygonal shape.

3. The spatial transcriptome biochip of claim 2, wherein edges and/or corners of the micro-cells are notched to form an orientation mark.

4. The spatial transcriptome biochip of claim 2, wherein the distance between adjacent ones of the microlocations is between 0 microns and 40 microns.

5. The spatial transcriptome biochip of any one of claims 1 to 4, wherein the diameter of the microwell structure ranges from 0.1 micron to 10 microns, and the center point distance between two adjacent microwells ranges from 0.1 micron to 20 microns.

6. The spatial transcriptome biochip according to any one of claims 1 to 4, wherein the long side of the transparent substrate has a value in the range of 10-100mm, and the short side has a value in the range of 5-50mm.

7. The spatial transcriptome biochip of any one of claims 1 to 4, wherein the size of the subregions ranges from 9mm x 9mm to 1875mm x 1875 mm.

8. The spatial transcriptome biochip of any one of claims 1 to 4, wherein the transparent substrate is one of glass, quartz, plastic, magnesium chloride, and gallium arsenide.

9. The spatial transcriptome biochip of any one of claims 1 to 4, wherein the microporous structure comprises a flared portion on a surface of the transparent substrate and a constricted portion inside the transparent substrate, the flared portion and the constricted portion communicating along a depth direction of the transparent substrate to form the microporous structure on the transparent substrate.

10. The spatial transcriptome biochip of any one of claims 1 to 4, wherein the transparent substrate has a first surface and a second surface, and the first surface and the second surface form a micropore region by photolithography and etching, respectively;

wherein the first surface and the second surface are two opposite surfaces of the transparent substrate.

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