A frozen section immunohistochemical staining machine

Abstract

The present invention relates to a frozen section immunohistochemical staining machine and method. A frozen section immunohistochemical staining machine has N (N≥2) staining chambers, and each staining chamber includes a reagent bottle assembly, a slide assembly, a sampling needle assembly, a robotic arm, and a heating assembly; each staining chamber can independently complete the immunohistochemical staining work. Frozen sections of an organ (tissue) are assembled into a section assembly, and reagent bottles matching the detection molecules are assembled into a reagent bottle assembly, and a group of sections are sampled and detected simultaneously, shortening the time of the entire experimental process. Through the frozen section staining machine and the immunohistochemical staining method relying on this staining machine, rapid immunohistochemical staining can be performed in a timely manner on the frozen sections of specimens delivered successively, thereby improving the accuracy of frozen section pathological diagnosis.

Description

Technical Field

[0001] The present invention relates to the technical field of medical devices, and particularly to a frozen section immunohistochemical staining machine. Background Art

[0002] Intraoperative rapid frozen section examination is an urgent consultation requested by a surgeon from a pathologist during the operation regarding the disease diagnosis related to the operation plan. It is the most important and difficult work in clinical pathology, featuring high technology, high difficulty, and high risk. The main reasons are the urgent time, as the doctor needs to issue a report within 30 minutes, and another important reason is the lack of technical support such as immunohistochemical staining in frozen section diagnosis.

[0003] Existing immunohistochemical staining equipment and methods cannot perform rapid immunohistochemical staining on frozen sections, thus making a more accurate diagnosis of frozen sections.

[0004] The reasons include: an immunohistochemical staining machine has only one staining functional area and cannot detect specimens submitted for examination successively in a timely manner on the same device; the robotic arm moves in a way with X / Y / Z degrees of freedom, resulting in a long action time; usually, a single sampling needle is used to sample each section successively, and the same molecules to be detected on different slides are detected using the reagent in the same reagent bottle, leading to a long test process cycle; the installation of reagent bottles and the placement of slides are not optimized into modules that are convenient for installation / dismantling, which is time-consuming and inconvenient during use; the glass slides have only one fixed position mode in the staining machine and cannot meet the needs of the two modes of sampling reaction and washing in the actual staining process, resulting in low efficiency of reaction and washing, prolonged time, and decreased quality; in the staining method of an automatic immunohistochemical staining machine, the catalytic enzyme is labeled on the secondary antibody. During detection, after the primary antibody recognizes and binds to the antigen, the secondary antibody binds to the primary antibody, and the catalytic enzyme on the secondary antibody catalyzes the substrate to develop color. Using this "two-step method" for immunohistochemical staining prolongs the immunohistochemical staining time. These factors result in a long full-process cycle and low efficiency of existing automatic immunohistochemical staining machines, making it impossible to apply the automated immunohistochemical staining technology in frozen section diagnosis. Summary of the Invention

[0005] The object of the present invention is to provide a frozen immunohistochemical staining machine and method. It is mainly a device and method capable of performing rapid immunohistochemical staining on clinical frozen sections, thereby improving the quality of clinical frozen section diagnosis and reducing the difficulty of diagnosis and the risk of misdiagnosis.

[0006] The technical solution of the present invention is a frozen section immunohistochemical staining machine, including a frozen section immunohistochemical staining machine and a method for immunohistochemical staining of frozen sections based on this staining machine; the frozen section immunohistochemical staining machine includes a main body and a staining chamber, a reagent bottle assembly, a slide assembly, a sampling needle assembly, a robotic arm, a temperature control assembly, a confluence pool, a liquid path assembly, a storage box, a refrigerator, and a desktop host arranged on the main body; the detection reagent in the reagent bottle assembly is dropped onto the glass slide of the slide assembly through the sampling needle assembly to stain the frozen section. During the staining process, the temperature control assembly heats and keeps the glass slide warm, and the finally stained waste liquid flows into the confluence pool; the staining chamber is located in the upper part of the main body, the confluence pool is located in the middle, the storage box and the refrigerator are located in the lower part, and the host is located in the upper left part;

[0007] There are more than 2 staining chambers on a frozen section immunohistochemical staining machine, which can be physically isolated or functionally partitioned. Each staining chamber contains an independent reagent bottle assembly, a slide assembly, a sampling needle assembly, a robotic arm, and a heating assembly. The sampling robotic arm is located in the upper part of the staining chamber, the reagent bottle assembly and the sampling needle assembly are located in the middle, and the heating assembly, the slide assembly, and the mode conversion robotic arm are arranged in the lower part. The slide assembly is directly placed on the heating assembly, and each staining chamber has the function of independently completing immunohistochemical staining. Among them, the reagent bottle assembly includes a reagent bottle rack and reagent bottles. The reagent bottle rack is a cuboid box structure for fixing a reagent bottle group. There are round holes at the bottom corresponding to the bottom of the reagent bottle, which is the position where the robotic arm presses the reagent bottle to squeeze out the detection reagent. There is a fixed insertion plate at the top for fixing the reagent bottle. The reagent bottle rack is provided with liquid level detection holes on both sides of the bottle and a hand-held position on the side for the loading and unloading of the reagent bottle assembly; there is a bottle mouth quantitative dispenser at the reagent bottle mouth and a sealing cap on the drip mouth; the slide module includes a glass slide support, a fixing plate, a flow limiter, and an electromagnetic switch, which are used to support and fix the glass slide and limit the added reagent to the area of the specimen to be tested. The electromagnetic switch is controlled by current to meet the needs of mode conversion. The slide assembly has a sampling reaction mode and a washing mode, and has the functions of horizontal / tilt position conversion and shaking to mix the liquid; the reagent bottle assembly and the slides are adapted to all staining chambers.

[0008] The sampling needle assembly includes a sampling needle, a sampling needle support, a developer bottle box, etc. At the working position corresponding to one glass slide (specimen to be tested), there are multiple sampling needles for independently dropping the detection reagent on each section.

[0009] Large reagent bottles and waste liquid buckets are placed in the storage box for storing fixatives, buffers, and collecting waste liquid. There is a liquid level detector on the storage bottle to ensure that the reagent is not too little and the waste liquid does not overflow too much.

[0010] The refrigerator is used to store the reagent bottle assembly.

[0011] The liquid path system includes a sample adding liquid path and a liquid discharging path. The inlet end of the sample adding liquid path is connected to a fixing liquid reagent bottle, a buffer reagent bottle, and a chromogenic reagent bottle. After passing through a diaphragm liquid pump and a pressure reducing valve, the outlet end branches and is connected to a sample adding needle in the staining chamber. The inlet end of the liquid discharging path is connected to a confluence pool, and the outlet end is connected to a waste liquid bottle, which collects the waste liquid into the waste liquid bottle by relying on the natural flow method.

[0012] The robotic arm assembly includes a sample adding robotic arm and a mode conversion robotic arm, which are respectively arranged at the top and bottom of the staining chamber and are composed of a transmission mechanism and a driving motor. The sample adding robotic arm is located above the reagent bottle assembly and acts on the reagent bottles in the reagent bottle assembly to drip the reagents in the reagent bottles onto the samples to be tested. The mode conversion robotic arm is located below the slide assembly near the slide holding position and acts on the slide assembly to change the position of the slide to meet the requirements of two different modes, namely the sample adding reaction and washing.

[0013] The desktop host controls the robotic arm, temperature control component, liquid path component, etc. through digital-to-analog / analog-to-digital conversion; the desktop host precisely controls the experimental process of the staining device through the immunohistochemistry experiment process module.

[0014] The method is a method for quickly performing immunohistochemical staining on frozen sections by relying on this frozen section immunohistochemical staining machine, including: grouping the prepared frozen sections from specimens of one organ (tissue) source and performing sample adding and staining simultaneously in one staining chamber; preferably, labeling the catalytic enzyme on the first antibody and using the "one-step method" for staining to achieve the purpose of quickly performing immunohistochemical staining on the specimens delivered successively and meeting the clinical requirements for pathological diagnosis of frozen sections within a short time.

[0015] The beneficial effects of the present invention are: (1) Through this frozen section immunohistochemical staining machine and the optimized immunohistochemical staining method, the immunohistochemical staining of frozen sections can be completed within 20 minutes, meeting the technical specification requirements for clinicians to make a frozen section pathological diagnosis within 30 minutes.

[0016] (2) There are multiple independent immunohistochemical staining chambers on one frozen immunohistochemical staining machine, which can meet the requirements for clinically frozen detection specimens to be sent for inspection one by one and quickly make a clinical pathological diagnosis.

[0017] (3) Grouping the frozen sections of one organ (tissue) and respectively and simultaneously adding reagents for detection in one staining chamber greatly shortens the experimental process time.

[0018] (4) Installing a set of commonly used (preset) antibody reagents for one organ (tissue) on one reagent bottle assembly is convenient for quickly and conveniently installing them into the staining chamber for detection and unloading and storing them after use.

[0019] (5) A frozen section of an organ (tissue) is placed on a slide assembly to form a group, and can be easily placed at a position matching the antibody in the reagent bottle assembly in the staining chamber for detection.

[0020] (6) The reagent bottle assembly and the slide assembly are adapted to all staining chambers and are more in number than the staining chambers, which can meet the need of adapting to the change of multiple samples of the same kind of organ (tissue) or different kinds of organ (tissue) combinations submitted for inspection in a short time.

[0021] (7) The slide assembly is provided with a sample adding reaction mode and a washing mode, and the position and angle of the slide are changed when reacting and washing the sample to accelerate the detection reaction speed and improve the washing efficiency.

[0022] (8) On each staining station corresponding to each slide (test sample), a plurality of sample adding needles are provided, and each sample adding needle is fixedly used to drop one kind of reagent. In this way, reagents can be quickly dropped on all the sections at the same time, shortening the sample adding time.

[0023] (9) For the immunohistochemical staining method carried out based on this frozen section immunohistochemical staining machine, it is preferred to label the catalytic enzyme on the first antibody and adopt the "one-step method" for staining to shorten the immunohistochemical staining time. Description of the Drawings

[0024] Figure 1 It is a schematic structural diagram of a frozen section immunohistochemical staining machine of the present invention.

[0025] Figure 2 It is a schematic structural diagram of the staining chamber of the present invention.

[0026] Figure 3 It is a schematic structural diagram of the reagent bottle assembly of the present invention.

[0027] Figure 4 It is a schematic structural diagram of the slide assembly of the present invention.

[0028] Figure 5 It is a schematic structural diagram of the robotic arm of the present invention.

[0029] Figure 6 It is a schematic structural diagram of the sample adding needle assembly of the present invention.

[0030] Figure 7 It is a schematic connection diagram of the liquid path system of the present invention.

[0031] Figure 8 It is a flow chart of the staining method of the present invention. Detailed Embodiments

[0032] To make the objectives, technical solutions and advantages of the present invention more clear, it should be noted that the orientation or positional relationships indicated by the terms "upper", "lower", "left", "right", "inner", "outer", etc. are based on the orientation or positional relationships shown in the drawings. These are only for convenience of description and simplification, rather than indicating or implying that the devices or components referred to must have a specific orientation. In addition, in the description of the present invention, the terms "installation" and "connection" should be understood in a broad sense.

[0033] The technical solutions of the present invention will be further described in detail below with reference to the accompanying drawings of the specification and embodiments.

[0034] As Figure 1 shown, a frozen section immunohistochemical staining machine of the present invention includes a main body 1 and a

[0035] staining chamber 2, a reagent bottle assembly 3, a slide assembly 4, a robotic arm 5, a heating assembly 6, a mainframe 7, a storage box 8, a refrigeration box 9 and a confluence pool 10. The main body 1 is of a box structure. The staining chamber 2 is located in the upper part of the main body 1, the confluence pool 10 is located in the middle, the storage box 8 and the

[0036] refrigeration box 9 are located in the lower part, and the desktop mainframe 7 is located in the upper left part of the main body 1. There are more than 2 staining chambers 2 on a frozen section immunohistochemical staining machine, which can be physically isolated or functionally partitioned. In this example, a frozen section immunohistochemical staining machine with 5 staining chambers 2 is used for illustration.

[0037] As Figure 2 shown, each staining chamber includes an independent reagent bottle assembly 3, a slide assembly 4, a robotic arm 5, a sampling needle assembly 21, and a heating assembly 6. The robotic arm 5 is located in the upper part of the staining chamber 2, the reagent bottle assembly 3 and the sampling needle assembly 21 are located in the middle, and the lower part is the heating assembly 6 and the slide assembly 4. The slide assembly 4 is directly placed on the heating assembly 6. Each staining chamber has the function of independently completing immunohistochemical staining.

[0038] As Figure 3 shown, the reagent bottle assembly 3 includes a reagent bottle 31, a fixed plug board 32 and a reagent bottle rack 33. The reagent bottle rack 33 is of a cuboid box structure and is used to fix a reagent bottle group. A hand-held position 331 is provided on the side of the reagent bottle rack, a round hole 332 corresponding to the reagent bottle mouth is provided at the top, and a round hole 333 corresponding to the reagent bottle bottom is provided at the bottom, which is the position where the robotic arm 5 presses the reagent bottle to extrude the detection reagent. A bottle mouth metering dispenser 311 is provided at the mouth of the reagent bottle 31.

[0039] As Figure 4As shown in the figure, the slide component 4 includes a fixed pressing plate 41, a slide holder 42, a flow limiter 411, and an electromagnetic switch 412, which are used to support and fix the slide 43 and confine the reagent after sample addition to the area to be tested. The electromagnetic switch 412 meets the needs of mode conversion. The slide component 4 has a sample addition reaction mode and a rinsing mode, and has functions of horizontal / tilt position conversion and shaking to mix the liquid. One side of the slide holder 42 is provided with a hand-held position 421 and a fixed buckle 422.

[0040] As Figure 5 shown in the figure, the robotic arm 5 includes a sample addition robotic arm 51, a mode conversion robotic arm 52, a moving stage 53, a guide rail 54, a drive motor 55, a synchronous pulley 56, a driven pulley 57, and a synchronous belt 58. The sample addition robotic arm 51 acts on the reagent bottle 31, and the mode conversion robotic arm 52 acts on the slide component 4.

[0041] As Figure 6 shown in the figure, the sample addition needle assembly 21 includes a liquid path system support 211, a chromogenic agent liquid pump 212, a sample addition needle holder 213, a sample addition needle 214, a chromogenic agent bottle box 215, and a chromogenic agent bottle 216. At the working station corresponding to one slide 43 (sample to be tested), multiple sample addition needles 214 are provided to independently drip reagents on each section.

[0042] As Figure 7 shown in the figure, the liquid path system is divided into a sample addition liquid path and a drainage liquid path. The inlet ends of the sample addition liquid path are respectively connected to the fixing liquid bottle 81, the buffer solution bottle 82, and the chromogenic agent bottle 216, and the outlet ends are branched and respectively connected to the sample addition needles 214 in the staining square 2, among which a diaphragm liquid pump 01 and a pressure reducing valve 02 are connected; the inlet end of the drainage liquid path is connected to the confluence pool 10, and the outlet end is connected to the waste liquid bottle 83, and the waste liquid is collected into the waste liquid bottle 83 by relying on the natural flow method.

[0043] As Figure 8 shown in the figure, the staining method flow chart includes: installation of the reagent bottle assembly 3 S1, installation of the slide component 4 S2, inserting the reagent bottle assembly 3 and the slide component 4 into the staining square bin 2 S3, fixing the specimen S4, washing S5, dripping the antibody S6, washing S7, developing color S8, washing S9, taking out the slide component 4 S10.

[0044] The working process of the frozen section immunohistochemical staining machine is described in detail below to further illustrate the technical solution of the present invention. The working process of the fully automatic immunohistochemical system includes the following steps.

[0045] 1. Installation of the reagent bottle assembly 3: Put the reagent bottle 31 for detecting 1 organ (tissue) marker molecule into the reagent bottle rack 33, assemble it into the reagent bottle assembly 3, and store it in a 4°C refrigerator. When in use, insert it with the bottle mouth downward into the working station in the staining square bin 2.

[0046] 2. Installation of the slide assembly 4: One type of organ (tissue) specimen submitted for clinical examination is made into a frozen section and placed on a slide holder 42. The fixing pressure plate 41 is covered, and the slide assembly 4 is assembled and inserted into the working station in the staining chamber 2. The biomarker detected on the specimen on the slide 43 is consistent with the type of antibody in the reagent bottle assembly 3.

[0047] 3. Tissue fixation: At the start of the experiment, the liquid path system aspirates the fixing solution bottle 81 in the storage box 8 and simultaneously drips it onto the slide 43 of the slide assembly 4 to fix the test specimen. At this time, the slide assembly 4 is in the detection position and remains in a horizontal position.

[0048] 4. Washing: The mode conversion robotic arm 52 converts the slide assembly 4 to the washing position. The electromagnetic switch 412 is turned on, and the slide 43 is tilted approximately 30 degrees. The fixing solution flows out, and then the liquid path system drips the buffer solution onto the slide 43 to wash the specimen.

[0049] 5. Antigen-antibody reaction: After washing is completed, the mode conversion robotic arm 52 converts the slide assembly 4 to the detection position. At this time, the slide is in a horizontal position, the electromagnetic switch 412 is turned off, and the sample adding robotic arm 51 moves downward and contacts the bottom of the reagent bottle 31.

[0050] And it pushes the reagent bottle 31 downward to drip the antibody in the reagent bottle 31 onto the sample on the slide 43 for the antigen-antibody reaction.

[0051] 6. Washing: After the reaction is completed, the mode conversion robotic arm 52 converts the slide assembly 4 to the detection position. The slide 43 is tilted approximately 30 degrees, the electromagnetic switch 412 is turned on, and the antibody reagent solution flows out. Then the liquid path system drips the buffer solution onto the slide 43 to wash the specimen.

[0052] 7. Chromogenic reaction: After washing is completed, the mode conversion robotic arm 52 converts the slide assembly 4 to the detection position. The slide 43 is in a horizontal position, and the liquid path system drips the chromogenic agent in the chromogenic agent bottle 216 onto the sample on the slide 43 for the chromogenic reaction.

[0053] 8. Washing: The mode conversion robotic arm 52 converts the slide assembly 4 to the washing position. The electromagnetic switch 412 is turned on, and the slide 43 is tilted approximately 30 degrees. The chromogenic agent flows out, and then the liquid path system drips the buffer solution onto the slide 43 to wash the specimen.

[0054] The entire process is precisely controlled by the mainframe 7.

[0055] It should be noted that the above description of the specific embodiments of the present invention is only for explaining the technical route and characteristics of the present invention, and its purpose is to enable those skilled in the art to understand the content of the present invention and implement it accordingly. However, the present invention is not limited to the above specific embodiments. Any changes or modifications made within the scope of the claims of the present invention should be covered by the protection scope of the present invention.

Claims

1. A frozen section immunohistochemical staining machine, characterized in that, An immunohistochemical staining machine for frozen sections and a method for immunohistochemical staining of frozen sections based on this staining machine; The immunohistochemical staining machine for frozen sections includes a main body, and a staining chamber, a reagent bottle assembly, a slide assembly, a sampling needle assembly, a robotic arm, a temperature control assembly, a confluence pool, a liquid path assembly, a storage box, a refrigerator, and a desktop host arranged on the main body. The staining chamber is located at the upper part of the main body, the confluence pool is located in the middle, the storage box and the refrigerator are located at the lower part, and the host is located at the upper left; The staining chamber includes a reagent bottle assembly, a slide assembly, a sampling needle assembly, a robotic arm, and a heating assembly; The reagent bottle assembly is characterized by including a reagent bottle rack and reagent bottles. A hand-held position is provided on the reagent bottle rack, and a quantitative dispenser for the bottle mouth is provided at the reagent bottle mouth; The slide assembly is characterized by including a glass slide support, a fixed pressing plate, a flow limiter, and a magnetic switch, which is used to support and fix the glass slide, limit the added reagent to the area of the specimen to be tested, and has a mode conversion function; The sampling needle assembly is characterized in that an independent sampling needle is equipped at each staining station of each slide; The robotic arm is characterized by including a sampling robotic arm, a mode conversion robotic arm, a moving stage, a guide rail, a driving motor, a synchronous pulley, a driven pulley, and a synchronous belt. The sampling robotic arm acts on the reagent bottle, and the mode conversion robotic arm acts on the slide assembly The method is characterized in that it is a method for quickly performing immunohistochemical staining on frozen sections relying on this immunohistochemical staining machine for frozen sections.

2. The frozen section immunohistochemical staining machine according to claim 1, wherein There are more than 2 staining chambers on one device, including two methods: (1) Physical isolation area; (2) Functional partition.

3. A frozen section immunohistochemical staining machine according to claim 1, characterized in that, The bottom of the staining chamber is a heating assembly and a slide assembly, above which are a reagent bottle assembly and a sampling needle assembly. The sampling robotic arm is located at the top, and the mode conversion robotic arm is located below the hand-held position of the glass slide.

4. A frozen section immunohistochemical staining machine according to claim 1, wherein, Each staining chamber has the function of independently completing immunohistochemical staining.

5. A frozen section immunohistochemical staining machine according to claim 1, characterized in that, The reagent bottle assembly assembles multiple reagent bottles into a reagent bottle group, and can simultaneously drip detection reagents onto different samples to be tested.

6. The cryostat immunohistochemical staining machine according to claim 1, wherein The slide assembly forms a glass slide group with the sections of one kind of tissue, and installs the matching reagent bottle assembly for simultaneous sampling and detection according to the marker molecule to be detected.

7. A frozen section immunohistochemical staining machine according to claim 1, wherein, The slide assembly is provided with a flow limiter and an electromagnetic switch, which can realize the conversion between the sampling reaction mode and the rinsing mode, and has the function of horizontal position / tilt position conversion, improving the reaction and rinsing efficiency.

8. A frozen section immunohistochemical staining machine according to claim 1, wherein, At each section station, a group of sampling needles is equipped, and each sampling needle is fixed to drip a kind of detection reagent, realizing rapid sampling and staining of a group of sections at the same time.

9. The cryostat immunohistochemical staining machine according to claim 1, wherein The reagent bottle assembly and the slide are adapted to all staining chambers, and are convenient and quick to install and disassemble.

10. A frozen section immunohistochemical staining machine according to claim 1, characterized in that, The catalytic enzyme is directly labeled on the first antibody, and the staining is carried out by a one-step method.

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