Method of manufacturing a sample fluid containing device and three-dimensional microscopic imaging method

Abstract

The application discloses a manufacturing method of a sample liquid containing device and a three-dimensional microscopic imaging method, and relates to the technical field of medical microscopic detection. The manufacturing method comprises the following steps: manufacturing a transparent base material into a column, wherein the column comprises two mutually perpendicular adjacent side surfaces; forming a through groove on the column, wherein the through groove extends along the length direction of the column and penetrates through the two parallel bottom surfaces of the column, the through groove occupies a right-angle side edge of the column and penetrates through the two mutually perpendicular adjacent side surfaces of the column; covering a cover glass on the through groove, wherein the space surrounded by the cover glass and the through groove forms a sample liquid containing groove; and making at least one end of the cover glass in the length direction be staggered from the end of the bottom surface of the column by a predetermined distance to form a sample liquid suction inlet. The sample liquid containing groove allows the liquid-based cells to be detected to pass through and can generate sufficient capillary force to suck in the sample liquid. The application enables the microscopic camera device to shoot liquid-based cell images from different angles and then combine the images into a real three-dimensional microscopic image.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of medical microscopic detection technology, in particular to a manufacturing method of a sample liquid containing device and a three-dimensional microscopic imaging method. BACKGROUND

[0002] With the development of image processing and digital imaging technology, liquid-based cells can be scanned and imaged by using digital microscopic imaging technology to obtain digital images. This enables pathologists or doctors to perform remote reading based on the digital images, thereby making accurate diagnoses on difficult cases. Two-dimensional microscopic imaging has gradually failed to meet the requirements of medical practice. In order to increase the amount of information of cell images and improve the diagnostic accuracy of doctors, three-dimensional imaging of liquid-based cell samples is often required.

[0003] The existing three-dimensional imaging technology often uses a microscopic camera to take pictures in X, Y and Z dimensions of a thin sheet device composed of a traditional glass slide and a cover glass. Different focal length images are taken in the depth direction (Z axis), and then combined into a three-dimensional image. However, the three-dimensional image obtained in this way is a pseudo three-dimensional image, because it can only provide three-dimensional information of different depths at a single angle, and cannot provide three-dimensional information of different depths at different angles. This pseudo three-dimensional image has application defects in the identification of some special liquid-based cells. For example, some cancer cells, the pseudo three-dimensional image cannot identify the characteristics of the lateral angle of the liquid-based cells, such as cell lateral defects and overlapping, which is of great significance to identify whether the sample liquid-based cells are benign or malignant.

[0004] Since the traditional glass slide and cover glass constitute a thin sheet device that can only take pictures at a vertical angle to obtain two-dimensional image data at a vertical angle, it is impossible to achieve three-dimensional imaging at multiple angles. Therefore, the existing technology based on the traditional thin sheet device cannot achieve real three-dimensional microscopic imaging of liquid-based cells. SUMMARY

[0005] In view of the above-mentioned defects or shortcomings in the prior art, the present application provides a manufacturing method of a sample liquid containing device and a three-dimensional microscopic imaging method of a sample liquid, which completely discards the traditional thin sheet structure composed of a glass slide and a cover glass, and creatively adopts a three-dimensional structure composed of a glass body and a cover glass. This enables the microscopic camera device to take pictures of liquid-based cell images from different angles, and then combine them into a real three-dimensional microscopic image. Doctors can obtain three-dimensional information of liquid-based cells at multiple angles through the three-dimensional microscopic image, which provides strong support for the diagnosis and decision-making of doctors.

[0006] In one aspect of the present application, a manufacturing method of a sample liquid containing device is provided, comprising the following steps:

[0007] The transparent substrate is made into a column-shaped carrier, the column includes at least two mutually perpendicular adjacent side surfaces; a through groove is formed on the carrier, the through groove extends along the length direction of the column and penetrates the two parallel bottom surfaces of the column, and the through groove occupies one straight corner side edge of the column and penetrates the two mutually perpendicular adjacent side surfaces of the column; a cover glass is fixed on the carrier and covers the two mutually perpendicular adjacent side surfaces of the column, and the space surrounded by the cover glass and the through groove forms a sample liquid containing groove; at least one end of the cover glass in the length direction is staggered from the end of the bottom surface of the carrier by a predetermined distance, thereby forming a sample liquid suction port of the sample liquid containing groove; the size of the sample liquid containing groove is set to allow the detected liquid-based cells to pass through and to generate sufficient capillary force to suck in the sample liquid.

[0008] Further, the material of the carrier and the cover glass is glass or quartz.

[0009] Further, the column is a cuboid; the length of the cuboid is set to 5-10 cm, the bottom surface of the cuboid is set to a square, and the side length of the square is set to 0.5-0.7 cm; the cross section of the sample liquid containing groove is set to a square, and the side length of the square is set to 20-30 μm.

[0010] Further, the cover glass is fixed on the carrier in a detachable manner.

[0011] Further, a buckle or a clamping groove is arranged on the cover glass, and a corresponding clamping groove or buckle is arranged on the carrier, and the cover glass is fixed on the carrier in a buckle connection manner.

[0012] Further, a first cover glass covers the first side surface of the column penetrated by the through groove, and a second cover glass covers the second side surface of the column penetrated by the through groove, the first side surface and the second side surface are adjacent and perpendicular, and at the intersection of the first cover glass and the second cover glass, the first cover glass completely covers the side surface of the second cover glass.

[0013] Another aspect of the present application also provides a three-dimensional microscopic imaging method of a sample liquid, including the following steps:

[0014] The sample liquid containing device is obtained according to the manufacturing method of the sample liquid containing device;

[0015] The light source irradiates the sample liquid containing groove through the carrier;

[0016] The first camera is aligned with the first outer side surface of the sample liquid containing groove, and the second camera is aligned with the second outer side surface of the sample liquid containing groove, the first outer side surface and the second outer side surface are adjacent and perpendicular, and the first camera and the second camera are located on the same plane and perpendicular to each other.

[0017] controlling the first camera and the second camera to move from one end of the sample liquid containing groove to the other end at a predetermined shooting frequency and moving speed;

[0018] sending the sample liquid image data acquired by the first camera and the second camera to an upper computer, and synthesizing a three-dimensional microscopic image by the upper computer.

[0019] Further, the column body is a cuboid, the first light source is vertically aligned with the first side of the slide body which does not expose the through groove, and the second light source is vertically aligned with the second side of the slide body which does not expose the through groove.

[0020] Further, the column body is a cuboid, and the light source irradiates the sample liquid containing groove along the opposite edges of the through groove of the slide body.

[0021] Further, the opposite edges of the through groove of the slide body are cut, and the cutting surface is perpendicular to the incident light of the light source.

[0022] The sample liquid containing device manufacturing method and three-dimensional microscopic imaging method have the following beneficial effects: the traditional slide plane structure is replaced by the three-dimensional structure of the slide body, so that the imaging device can shoot the image of the liquid-based cell at different angles, and then the real three-dimensional microscopic image is conveniently synthesized; the sample liquid containing groove is arranged on the slide body in a structure, so that the microscopic imaging device is not affected by the non-smooth surface of the etched groove or the laser cutting groove when shooting, and clearer image data can be obtained; the slide body, the cover glass and the through groove form the sample liquid containing groove with a capillary structure, which can be automatically filled with sample liquid by capillary force when the sample liquid is filled, which is very convenient; two orthogonal microscopic cameras shoot the sample liquid containing groove at a predetermined shooting frequency and moving speed, and the shooting image is three-dimensionally synthesized, so that the real three-dimensional microscopic image with much more information than the traditional technology can be obtained. BRIEF DESCRIPTION OF DRAWINGS

[0023] Other features, objects and advantages of the present application will become more apparent from the following detailed description of non-limiting embodiments, made with reference to the accompanying drawings:

[0024] Figure 1 is a schematic diagram of a sample liquid containing device of a thin sheet type composed of a slide and a cover glass in the prior art;

[0025] Figure 2 is a flowchart of a sample liquid containing device manufacturing method according to an embodiment of the present application;

[0026] Figure 3 is a cross section of a sample liquid containing device according to an embodiment of the present applicationFigure 1 ;

[0027] Figure 4 is a sectional view of a sample liquid containing device provided by another embodiment of the present application Figure 2 ;

[0028] Figure 5 is a perspective view of a sample liquid containing device provided by another embodiment of the present application

[0029] Figure 6 is a schematic view of a three-dimensional microscopic imaging system provided by another embodiment of the present application Figure 1 ;

[0030] Figure 7 is a schematic view of a three-dimensional microscopic imaging system provided by another embodiment of the present application Figure 2 ;

[0031] Figure 8 is a flow chart of a method for three-dimensional microscopic imaging of a sample liquid provided by another embodiment of the present application

[0032] Reference signs:

[0033] 100 - thin slice sample liquid containing device; 101 - glass slide; 102 - cover glass; 103 - microscope objective; 104 - sample liquid; 200 - sample liquid containing device; 201 - glass body; 202 - cover glass; 203 - through groove; 204 - sample liquid containing groove; 205 - sample liquid suction port; 206 - clamping groove; 207 - clamping buckle; 300 - three-dimensional microscopic imaging system; 301 - first camera; 302 - second camera; 303 - first light source; 304 - second light source; 305 - first side surface; 306 - second side surface. DETAILED DESCRIPTION

[0034] In order to make the objects, technical solutions and advantages of the embodiments of the present application clearer, the technical solutions in the embodiments of the present application will be described clearly and completely below with reference to the drawings in the embodiments of the present application. Obviously, the described embodiments are only some of the embodiments of the present application, but not all of the embodiments of the present application. Based on the embodiments in the present application, all other embodiments obtained by those of ordinary skill in the art without creative work fall within the scope of protection of the present application.

[0035] The terms used in the embodiments of the present application are only for the purpose of describing the specific embodiments, and are not intended to limit the present application. The singular forms "a", "an" and "the" used in the embodiments of the present application and the appended claims are also intended to include the plural forms, unless the context clearly indicates otherwise.

[0036] It should be understood that, although the terms first, second, third, etc. can be employed in describing the acquisition modules of the embodiments of the present application, these acquisition modules should not be limited by these terms. These terms are only used to distinguish one acquisition module from another.

[0037] Depending on the context, the word "if" as used herein can be interpreted to mean "when" or "while" or "in response to determining" or "in response to detecting." Similarly, the phrase "if it is determined" or "if [a stated condition or event] is detected" can be interpreted to mean "upon determining" or "in response to determining" or "upon detecting [the stated condition or event]" or "in response to detecting [the stated condition or event]."

[0038] It should be noted that the terms "upper", "lower", "left", "right", and the like as described in the embodiments of the present application are described in the angle shown in the drawings, and should not be understood as a limitation on the embodiments of the present application. In addition, in the context, it should also be understood that when referring to an element formed "on" or "under" another element, it can be directly formed "on" or "under" another element, or indirectly formed "on" or "under" another element through an intermediate element.

[0039] Figure 1 A slice sample liquid containing device 100 formed by a glass slide 101 and a cover glass 102 is shown. The glass slide 101 refers to a carrier glass, and the cover glass 102 refers to a cover glass. The materials of the glass slide and the cover glass are glass or quartz. Generally, the cover glass 102 is arranged on the side of the glass slide 101 close to the microscope objective 103, and the sample liquid 104 is arranged between the glass slide 101 and the cover glass 102. This conventional slice sample liquid containing device 100 can only allow the microscope lens or the microscopic camera device to observe or shoot vertically from above the cover glass 102 downward. Although different focal depth samples can be shot, the image of the liquid-based cell in the sample liquid cannot be obtained from other angles. Therefore, for the slice sample liquid containing device 100, the multi-angle image of the liquid-based cell in the sample liquid cannot be obtained, and a real three-dimensional image containing multi-angle information cannot be synthesized.

[0040] In order to make up for the defects of the conventional slice sample liquid containing device 100, referring to Figure 2 An embodiment of the present application provides a manufacturing method of a sample liquid containing device capable of obtaining a three-dimensional structure, thereby facilitating the acquisition of multi-angle image information of the sample liquid. The method comprises:

[0041] In step S101, a transparent substrate is made into a columnar glass carrier, and the columnar body includes two mutually perpendicular adjacent side surfaces.

[0042] The transparent substrate in the embodiment can be selected from one of glass or quartz, and is made into a columnar carrier. The carrier in the embodiment is a short form of carrier glass, and has the same use and material as the carrier slide in the prior art. However, the carrier slide in the prior art has a sheet structure, and the carrier in the embodiment is made into a columnar structure. The columnar body in the embodiment includes a cylinder, a prism or any other type of columnar body with the same bottom shape and parallel to each other. Meanwhile, the columnar body also includes two adjacent sides perpendicular to each other, for example, a cuboid, a cube, a right triangular prism, a right trapezoidal quadrangular prism and the like.

[0043] It should be noted that the columnar body is required to have two adjacent sides perpendicular to each other, because this makes the columnar body have at least one right-angled side edge, and the through groove is arranged at the right-angled side edge, so that the sample liquid in the through groove can be observed at two orthogonal angles on the two perpendicular sides at the same time, thereby more easily obtaining a three-dimensional composite image at different angles.

[0044] In step S102, a through groove is formed on the carrier by etching or laser cutting, the through groove extends along the length direction of the columnar body and penetrates the two parallel bottom surfaces of the columnar body, and the through groove occupies one right-angled side edge of the columnar body and penetrates the two adjacent sides perpendicular to each other of the columnar body.

[0045] Specifically, taking a cuboid with a square bottom as an example, the length of the cuboid is set to 5-10 cm, and the side length of the bottom surface of the cuboid is set to 0.5-0.7 cm. A through groove is formed on one side edge of the carrier in the shape of a cuboid by etching or laser cutting. Since the etching process and the laser cutting process are prior art in the field, the process of forming the through groove will not be described herein. The through groove extends along the length direction of the cuboid, penetrates the two parallel bottom surfaces of the cuboid, occupies one side edge of the cuboid, and penetrates the two adjacent sides perpendicular to each other of the cuboid. In other words, by digging away one side edge of the cuboid, a through groove is formed, which penetrates the two bottom surfaces of the cuboid and is parallel to the side edge of the cuboid. The through groove is preferably L-shaped in cross section, but can also be other shapes such as an arc-shaped cross section, which is not limited herein.

[0046] In step S103, a cover slide is fixed on the carrier and covers the two adjacent sides perpendicular to each other of the columnar body. The space surrounded by the cover slide and the through groove constitutes a sample liquid containing groove.

[0047] The length of the cuboid is set to 5-10 cm, and the length of the side of the bottom surface of the cuboid is set to 0.5-0.7 cm. In one embodiment, the cover glass can be a complete L-shaped cross-section sheet structure with a thickness of 0.1-0.2 mm, which is fixed on the glass slide by bonding to cover two adjacent sides of the cuboid perpendicular to each other, i.e. to cover the through groove. The space surrounded by the cover glass and the through groove constitutes the sample liquid containing groove. At this time, the sample liquid containing groove can be observed on both vertical cover glass sides. In another embodiment, the cover glass can be two independent sheet parts, which cover two adjacent sides of the cuboid exposing the through groove. Preferably, at the intersection of the two independent cover glasses, one cover glass completely covers the side of the other cover glass, so that a face contact is formed between the cover glasses, which is more airtight and can ensure that the sample liquid in the sample liquid containing groove does not overflow.

[0048] In order to facilitate disassembly and assembly, the cover glass can be fixed by bonding or other methods, or can be connected by a detachable buckle. For example, a clamping groove is provided on the side of the glass slide, and a buckle matching the clamping groove is provided on the cover glass. The buckle of the cover glass is aligned with the clamping groove of the glass slide and pressed, so that the cover glass is attached to the glass slide. The buckle structure makes the disassembly and assembly of the cover glass and the glass slide more convenient, thereby simplifying the manufacturing process.

[0049] In step S104, at least one end of the cover glass in the length direction is offset from the end of the bottom surface of the glass slide by a predetermined distance, thereby forming a sample liquid suction port of the sample liquid containing groove.

[0050] Specifically, at least one end of the cover glass in the length direction is offset from the end of the bottom surface of the glass slide by a predetermined distance to form a sample liquid suction port. The predetermined distance of the offset cannot be too large or too small, and is preferably 2-4 mm. This size is more conducive to quickly sucking in the sample liquid. A predetermined distance of less than 2 mm is not easy to suck in the sample liquid, and a predetermined distance of more than 4 mm wastes the effective space of the sample liquid containing groove.

[0051] In step S105, the size of the sample liquid containing groove is set to allow the detected liquid-based cells to pass through and to generate sufficient capillary force to suck in the sample liquid.

[0052] Specifically, the liquid-based cells in the sample liquid are taken as an example of cancer cells, and the size of the liquid-based cells is usually 10-20 μm. Therefore, the sample liquid containing groove should be able to contain the liquid-based cells to be detected, and the width or diameter of the sample liquid containing groove should not be less than 20 μm. At the same time, the sample liquid containing groove needs to use the capillary force generated by the capillary tube to suck the sample liquid dropped in, and therefore the diameter or width of the sample liquid containing groove should be able to generate a large enough capillary force. In combination with the above factors, it is verified through a large number of experiments that when the diameter or width of the sample liquid containing groove is set to 20-30 μm, most types of liquid-based cells can pass through, and a large enough capillary force can be generated.

[0053] The manufacturing method of the sample liquid containing device provided by the embodiment can obtain a three-dimensional sample liquid containing device, facilitate the two camera devices to capture images of the liquid-based cells at different angles, and further synthesize a real three-dimensional microscopic image. The sample liquid containing groove is arranged at the corner of the glass slide, so that the two microscopic camera devices are not affected by the uneven surface of the etched groove or the laser cutting groove when capturing images, and clearer image data can be obtained. The glass slide, the cover glass and the through groove form the sample liquid containing groove of the capillary structure, which can be automatically filled with sample liquid by capillary force when the sample liquid is injected, which is very convenient.

[0054] Referring to Figures 3-5 Another embodiment of the present application also provides a sample liquid containing device 200 obtained by the above manufacturing method, which comprises a glass slide 201 and a cover glass 202. The glass slide 201 is a column structure with a through groove 203 (the embodiment is taken as an example of a cuboid structure). The through groove 203 extends along the length direction of the cuboid and penetrates two bottom surfaces of the column. The through groove 203 occupies one right-angle edge of the cuboid and penetrates two adjacent side surfaces forming the right-angle edge. The cover glass 202 is fixed on the glass slide 201 and covers the two side surfaces of the through groove 203 in the length direction of the glass slide 201. The space surrounded by the through groove 203 and the cover glass 202 forms a sample liquid containing groove 204. At least one end of the cover glass 202 in the length direction is staggered from the end of the bottom surface of the glass slide 201 by a predetermined distance to form a sample liquid suction port 205 of the sample liquid containing groove 204. The size of the sample liquid containing groove 204 is set to allow the liquid-based cells to be detected to pass through and to generate a sufficient capillary force to suck the sample liquid.

[0055] It should be noted that the sample liquid containing groove 204 of the embodiment finally occupies one edge of the cuboid structure of the glass slide 201, that is, the right-angle edge of the cuboid is excavated to form the through groove 203, so that the sample liquid containing groove 204 is in an open state on two adjacent sides of the cuboid. The camera can directly take microphotographs at different angles through the cover glass 202 covering the two adjacent sides penetrated by the through groove 203, without the need to penetrate the glass slide 201 for shooting. The sample liquid containing groove 204 arranged in the above manner has the advantage that if an etched pit or a laser-cut groove is formed at other positions, the pit has only one exposed surface on the outer side of the column, or the groove is completely closed by the glass slide 201 and only the bottom surface is penetrated, which means that at most only one camera can shoot through the cover glass 202, and other cameras can only shoot through the etched surface or the laser-cut surface of the groove of the glass slide 201. However, the surface of the glass etched or laser-cut is not smooth, and shooting through the etched surface or the laser-cut surface will affect the shooting effect of the micro-camera, and further affect the final three-dimensional imaging effect. The cover glass 202 is a glass sheet made by special process, and has a smooth surface and high light transmittance. The arrangement of the sample liquid containing groove 204 of the embodiment can enable two cameras at different angles to directly shoot the sample liquid containing groove 204 through the cover glass 202, greatly improving the image quality of microphotography.

[0056] In another embodiment, the length of the cuboid of the sample liquid containing device 200 is set to 5-10 cm, the cross section and the bottom surface of the cuboid are square, and the side length of the square is set to 0.5-0.7 cm. Taking cancer cells as an example, the liquid-based cells are usually 10-20 μm in size, and the sample liquid containing groove 204 should be able to contain the detected liquid-based cells, so the width or diameter of the sample liquid containing groove 204 should not be less than 20 μm. At the same time, the sample liquid containing groove 204 needs to use the capillary force generated by the capillary tube to suck the sample liquid dropped in, so the diameter or width of the sample liquid containing groove 204 should ensure that a large enough capillary force is generated. Considering the above factors, the diameter or width of the sample liquid containing groove 204 is set to 20-30 μm through a large number of experiments, which can allow most types of liquid-based cells to pass through, and can also generate a large enough capillary force.

[0057] Further, the cover glass 202 is fixedly connected to the glass slide 201, for example, by adhesion.

[0058] Further, referring to Figures 3-4The coverslip 202 and the glass carrier 201 are connected by a detachable structure, for example, by a snap-fit ​​connection. Specifically, a slot 206 is provided on the side of the glass carrier 201, and a snap 207 matching the slot 206 is provided on the coverslip 202. By aligning the snap 207 of the coverslip 202 with the slot 206 on the glass carrier 201 and pressing it, the coverslip 202 can be pressed tightly against the glass carrier 201. This snap-fit ​​structure makes the assembly and disassembly of the coverslip 202 and the glass carrier 201 more convenient, thereby simplifying the manufacturing process.

[0059] Furthermore, participate Figure 3 The cover glass 202 can be an integral structure bent at a right angle, so that it can be installed as a whole on the glass carrier, covering the two adjacent sides of the through groove 203 exposed on the column.

[0060] Further, see Figure 4 The coverslip 202 can be two separate parts, each covering one of the two adjacent sides of the column exposed by the through groove 203. Preferably, at the intersection of the two separate coverslips 202, one coverslip 202 completely covers the side of the other coverslip 202, so that the coverslips form a surface contact, which makes the seal more tight and ensures that the sample liquid in the sample liquid receiving tank 204 will not overflow.

[0061] Further, see Figure 5 At least one end of the coverslip 202 is offset from the bottom end of the glass substrate 201 by a predetermined distance in the longitudinal direction to form a sample liquid aspiration port 205. The predetermined offset distance is preferably 2 to 4 mm, which is more conducive to rapid aspiration of sample liquid. A predetermined distance of less than 2 mm makes it difficult to aspirate sample liquid, while a predetermined distance of more than 4 mm wastes the effective space of the sample liquid receiving tank 204.

[0062] The sample liquid containing device 200 provided in this embodiment replaces the traditional thin sheet structure with a three-dimensional structure of a glass carrier, which facilitates two imaging devices to capture images of liquid-based cells from different angles, thereby synthesizing a realistic three-dimensional microscopic image. The sample liquid containing groove 204 is set at the corner of the glass carrier 201, so that the two microscopic imaging devices at different angles are not affected by the uneven surface of the etched groove or laser-cut groove when taking pictures, and can obtain clearer image data. The glass carrier 201, coverslip 202 and through groove 203 form a capillary structure sample liquid containing groove 204, which can be automatically filled with sample liquid by capillary force when injecting sample liquid, which is very convenient.

[0063] See Figures 6-7Another embodiment of the present application also provides a three-dimensional microscopic imaging system 300, comprising the sample liquid containing device 200 obtained according to the above manufacturing method embodiment, a microscopic camera device, a light source and a controller. Wherein the cover glass is in the shape of a cuboid.

[0064] The microscopic camera device comprises a first camera 301 and a second camera 302 vertically aligned to cover the two cover glasses 202 of the through groove 203 (i.e. vertically aligned to the two adjacent sides penetrated by the through groove), and the first camera 301 and the second camera 302 are located on the same plane and are arranged perpendicular to each other.

[0065] The light source is used to irradiate the sample liquid containing groove 204 through the cover glass 201. Referring to Figure 6 In an optional embodiment, the light source comprises a first light source 303 and a second light source 304, the first light source 303 is aligned to the first side 305 of the cover glass 201 which does not expose the through groove 203, and the second light source 304 is aligned to the second side 306 of the cover glass 201 which does not expose the through groove 203. Referring to Figure 7 In another optional embodiment, the light source irradiates the cover glass 201 along the opposite edges of the through groove 203 on the cover glass 201, the light is divided into two, respectively entering the cover glass 201 on both sides of the edge, and then providing light for the sample liquid containing groove 204. More preferably, the opposite edges of the through groove 203 on the cover glass 201 are cut off, and the cutting surface is perpendicular to the incident light of the light source, so that more light can be introduced in the case of a single light source or a group of light sources.

[0066] Further, the controller is used to control the first camera 301 and the second camera 302 to move from one end to the other end of the sample liquid containing groove 204 at a predetermined shooting frequency and moving speed, and send the obtained image data at two different angles to the upper computer, and the upper computer combines them into a three-dimensional image. More preferably, the controller moves the first camera 301 and the second camera 302 from one end of the cover glass 202 to the other end of the cover glass 202 when shooting. Only the covered area of the cover glass 202 is shot because: on the one hand, the cover glass 202 is made by deposition process, so the light transmittance is high and will not be affected by the uneven etching surface or the laser cutting surface of the through groove 203; on the other hand, the standard columnar containing space formed after the cover glass 202 covers the through groove 203 is full of sample liquid, and the imaging quality of the regular-shaped and bubble-free sample liquid is better.

[0067] Optionally, the controller can also be used to control the opening and closing of the light source.

[0068] Referring to Figure 8Another embodiment of the present application also provides a sample liquid three-dimensional microscopic imaging method based on a three-dimensional microscopic imaging system, comprising the following steps:

[0069] Step S201, manufacturing a sample liquid containing device according to the manufacturing method embodiment of the sample liquid containing device;

[0070] Step S202, allowing a light source to irradiate the sample liquid containing groove through the slide body;

[0071] Specifically, in an optional embodiment, the column is a cuboid, and the first light source is vertically aligned with the first side of the slide body which does not expose the through groove, and the second light source is vertically aligned with the second side of the slide body which does not expose the through groove. In another optional embodiment, the column is a cuboid, and the light source irradiates the sample liquid containing groove along the opposite edges of the through groove of the slide body, and the opposite edges of the through groove of the slide body are cut, and the cutting surface is perpendicular to the incident light of the light source.

[0072] Step S203, allowing a first camera to be aligned with the first outer side of the sample liquid containing groove, and allowing a second camera to be aligned with the second outer side of the sample liquid containing groove, the first outer side and the second outer side being adjacent and perpendicular, the first camera and the second camera being located on the same plane and being perpendicular to each other;

[0073] Step S204, controlling the first camera and the second camera to move from one end of the sample liquid containing groove to the other end at a predetermined shooting frequency and moving speed;

[0074] Step S205, sending the sample liquid image data obtained by the first camera and the second camera to an upper computer, and synthesizing a three-dimensional microscopic image by the upper computer.

[0075] The above description is only the preferred embodiment of the present application. Those skilled in the art should understand that the disclosure range involved in the present application is not limited to the technical solutions formed by the specific combination of the above technical features, and should also cover other technical solutions formed by any combination of the above technical features or their equivalent features without departing from the above disclosure concept. For example, the above features are replaced with the technical features disclosed in the present application (but not limited to) having similar functions to form technical solutions.

Claims

1. A method for manufacturing a sample liquid containing device for three-dimensional microscopic imaging, characterized in that, Includes the following steps: A transparent substrate is fabricated into a columnar glass carrier, the glass carrier comprising two adjacent perpendicular sides; A through groove is formed on the glass carrier, the through groove extends along the length of the column and penetrates two parallel bottom surfaces of the column, the through groove occupies one right-angled side edge of the column and penetrates two adjacent mutually perpendicular side surfaces of the column; A slot is provided on the coverslip, and a corresponding buckle is provided on the glass carrier. The coverslip is fixed to the glass carrier by the buckle connection and covers two adjacent perpendicular sides of the column. The space enclosed by the coverslip and the through groove constitutes the sample liquid receiving tank. The glass carrier and the coverslip are made of quartz. The coverslip is a complete L-shaped sheet structure with a thickness of 0.1-0.2 mm. At least one end of the coverslip along its length is offset from the bottom end of the glass substrate by a predetermined distance, thereby forming the sample liquid intake port of the sample liquid receiving tank; The sample solution container is sized to allow the liquid-based cells to be tested to pass through and to generate sufficient capillary force to draw in the sample solution.

2. The method for manufacturing a sample liquid containing device for three-dimensional microscopic imaging according to claim 1, characterized in that: The column is a cuboid; Set the length of the cuboid to 5-10cm, set the base of the cuboid to a square, and set the side length of the square to 0.5-0.7cm. The cross-section of the sample liquid container is set to be square, and the side length of the square is set to 20-30 mm.

3. A three-dimensional microscopic imaging method for a sample liquid, characterized in that, Includes the following steps: The sample liquid containing device for three-dimensional microscopic imaging is obtained by the manufacturing method of the sample liquid containing device according to claim 1 or 2; The light source illuminates the sample liquid container through the glass carrier; The first camera is aligned with the first outer side of the sample liquid container, and the second camera is aligned with the second outer side of the sample liquid container. The first outer side and the second outer side are adjacent to and perpendicular to each other. The first camera and the second camera are located on the same plane and are perpendicular to each other. The first and second cameras are controlled to move from one end of the sample liquid container to the other end at a predetermined shooting frequency and moving speed; The sample liquid image data acquired by the first and second cameras are sent to the host computer, and then synthesized into a three-dimensional microscopic image by the host computer.

4. The three-dimensional microscopic imaging method for a sample liquid according to claim 3, characterized in that, Also includes: The column is a cuboid; The first light source is vertically aligned with the first side of the glass carrier where the through groove is not exposed, and the second light source is vertically aligned with the second side of the glass carrier where the through groove is not exposed.

5. The three-dimensional microscopic imaging method for a sample liquid according to claim 3, characterized in that, Also includes: The column is a cuboid; The light source is directed to illuminate the sample liquid container along the opposite edges of the through groove in the glass substrate.

6. The three-dimensional microscopic imaging method for a sample liquid according to claim 5, characterized in that, Also includes: The edges opposite to the through groove of the glass carrier are cut off, and the cut surface is perpendicular to the incident light from the light source.

Images