CN111504739A - Immunohistochemical slide glass for detection - Google Patents
Immunohistochemical slide glass for detection Download PDFInfo
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- CN111504739A CN111504739A CN202010318770.6A CN202010318770A CN111504739A CN 111504739 A CN111504739 A CN 111504739A CN 202010318770 A CN202010318770 A CN 202010318770A CN 111504739 A CN111504739 A CN 111504739A
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- G—PHYSICS
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- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N1/30—Staining; Impregnating ; Fixation; Dehydration; Multistep processes for preparing samples of tissue, cell or nucleic acid material and the like for analysis
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- G—PHYSICS
- G02—OPTICS
- G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
- G02B21/00—Microscopes
- G02B21/34—Microscope slides, e.g. mounting specimens on microscope slides
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N1/2813—Producing thin layers of samples on a substrate, e.g. smearing, spinning-on
- G01N2001/2833—Collecting samples on a sticky, tacky, adhesive surface
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Abstract
The invention provides a glass slide for immunohistochemical detection, which at least comprises: a slide substrate at least partially covered with an adhesion layer; the glass slide is provided with a detection area, a control area and a label area, the control area is provided with a positive tissue chip, the positive tissue chip comprises a plurality of positive control tissues, the detection area and the control area are covered with an adhesion layer, and the positive tissue chip is positioned above the adhesion layer. The positive tissue chip can contain COVID-19 tissues positive to the novel coronavirus antigen. The method and the device can ensure that each slice has corresponding positive control tissues, the flaking process not only accords with a 15189 quality management system, but also shortens time, simplifies the process, improves efficiency and unifies standards. The anti-dropping tablet ensures strong adhesion of the positive control tissue at the early stage, ensures that the tablet does not drop in the later dyeing process, simultaneously ensures the antigen retention time of the positive control, ensures the antigen timeliness and prevents the positive control chip from degrading.
Description
Technical Field
The invention relates to the field of medical detection, in particular to a glass slide for immunohistochemical detection.
Background
The clinical detection needs to really achieve high sensitivity, strong specificity and short detection time, and is difficult due to the fact that the three methods need a key step except the characteristics of the experiment, namely the three methods need to separate the DNA/RNA or the antibody of the virus from the blood sample, but the scientific separation step can lead to low original blood sensitivity, partial or most of the virus is lost in the course of extraction, and the virus quantity is determined by whether the kit is scientific or not.
Then where is the best sample collection site? The lungs, including alveolar lavage, pharyngeal swabs, sputum smears, etc., without doubt; what is the most appropriate sample processing method? Detecting in situ without extraction. Therefore, cells collected by a throat swab are prepared into a tissue wax block, antigen loss is avoided to the maximum extent without an extraction method, and a new coronavirus antibody is detected in situ by an immunohistochemical method.
In order to avoid the generation of immunohistochemical false negative and false positive results as much as possible, the requirements of the quality control management system of the Chinese medical society and the implementation rules of the third-level hospital review standard (2011 edition) are specifically specified: positive control must be established for immunohistochemical staining to ensure stable and reliable staining results. The current general method for immunohistochemical positive control is to find biological tissues containing IgG/IGM antibodies (related to new coronavirus) as positive control, simultaneously fish the positive control tissues with the thickness of 5um and the section of the tissues to be detected on the same glass slide, prevent the tissues from falling off or enzyme degradation through a series of steps such as baking the glass slide, place the glass slide carrying the positive control tissues and the section of the tissues to be detected on the same condition for immunohistochemical reaction, and finally perform comparison and interpretation. The method is tedious, time-consuming and free of unified quality control standards, and the workload of a plurality of hospitals, particularly a hospital three, is large at present, so that positive control on each section is difficult to ensure, and an uncertain result is inevitably generated.
Disclosure of Invention
In view of the above-described drawbacks of the prior art, it is an object of the present invention to provide a slide glass for immunohistochemical detection. The glass slide is a chip glass slide with a positive control tissue, and can ensure that the positive control tissue required by various human diseases is covered at the same time so as to solve the practical problems encountered in clinical work.
In a first aspect, the present invention provides a slide for immunohistochemical detection, the slide comprising at least: a slide substrate at least partially covered with an adhesion layer; the glass slide is provided with a detection area, a control area and a label area, the control area is provided with a positive tissue chip, the positive tissue chip comprises a plurality of positive control tissues, the detection area and the control area are covered with an adhesion layer, and the positive tissue chip is positioned above the adhesion layer.
The second aspect of the present invention provides the use of the aforementioned slide glass for immunohistochemical detection in immunohistochemical detection.
The third aspect of the present invention provides a method for preparing the slide glass for immunohistochemical detection, comprising at least the steps of:
1) covering at least part of the surface of the glass slide substrate with an adhesive substance;
2) plating the positive tissue chip on the control area;
3) the surface of the positive tissue chip is covered with anti-degradation substances.
As described above, the slide glass for immunohistochemical detection of the present invention has the following advantageous effects:
can ensure that each section has corresponding positive control tissue, the flaking process not only accords with the 15189 quality management system, but also greatly shortens the time, simplifies the process, improves the efficiency and unifies the standard. The anti-dropping tablet ensures strong adhesion of the positive control tissue at the early stage, ensures that the tablet does not drop in the later dyeing process, simultaneously ensures the antigen retention time of the positive control, ensures the antigen timeliness and prevents the positive control chip from degrading.
Drawings
FIG. 1 is a schematic view showing the structure of a slide glass for immunohistochemical detection according to an embodiment of the present invention.
FIG. 2 is a side view of a slide for immunohistochemical detection according to one embodiment of the present invention.
FIG. 3 is a view showing a slide glass for immunohistochemical detection of the present invention stored at room temperature for half a year.
Description of the element reference numerals
1 glass slide substrate
2 adhesive layer
3 detection zone
4 control zone
41 positive tissue chip
42 degradation prevention layer
5 label area
51 chemical corrosion resistant coating
Detailed Description
The following description of the embodiments of the present invention is provided for illustrative purposes, and other advantages and effects of the present invention will become apparent to those skilled in the art from the present disclosure.
Please refer to fig. 1 to 3. It should be understood that the structures, ratios, sizes, and the like shown in the drawings and described in the specification are only used for matching with the disclosure of the specification, so as to be understood and read by those skilled in the art, and are not used to limit the conditions under which the present invention can be implemented, so that the present invention has no technical significance, and any structural modification, ratio relationship change, or size adjustment should still fall within the scope of the present invention without affecting the efficacy and the achievable purpose of the present invention. In addition, the terms "upper", "lower", "left", "right", "middle" and "one" used in the present specification are for clarity of description, and are not intended to limit the scope of the present invention, and the relative relationship between the terms and the terms is not to be construed as a scope of the present invention.
It should be noted that, for the sake of easy observation, the thickness ratios of the respective components in fig. 2 do not coincide with the actual ratios, and the main function of fig. 2 is to explain the relationship between the respective components.
As shown in fig. 1 and 2, an embodiment of the present invention provides a slide for immunohistochemical detection, the slide including at least: a slide substrate 1, as shown in fig. 2, said slide substrate 1 being at least partially covered with an adhesion layer 2; the glass slide is provided with a detection area 3, a control area 4 and a label area 5, the control area 4 is provided with a positive tissue chip 41, the positive tissue chip 41 comprises a plurality of positive control tissues, the detection area 3 and the control area 4 are covered with an adhesion layer 2, and the positive tissue chip 41 is positioned above the adhesion layer.
Alternatively, the detection zone 3, the control zone 4 and the label zone 5 are arranged along the length of the slide.
The detection area is used for adhering a target to be detected.
The main components of the glass slide substrate are silicon oxide and aluminum oxide, and the higher the content of silicon dioxide is, the better the chemical stability is; meanwhile, the less impurities are contained, and the higher the material density and the structural stability are.
Further, the adhesive layer 2 is composed of an adhesive substance selected from 3-Aminopropyl-3-ethoxysilane (3-Aminopropyl-triethoxy silane, APES), Poly-L-lysine (Poly-L-L ysine) or gelatin.
Preferably, Poly-L-lysine (Poly-L-L ysine).
In a preferred embodiment, the positive tissue chip 41 is covered with an anti-degradation layer 42.
Optionally, the degradation prevention layer 42 is composed of a degradation prevention substance selected from paraffin.
When an immunohistochemical experiment is carried out, the slide glass needs to be baked, and the degradation-resistant substance can fall off or melt, so that the use is not influenced.
In the positive tissue chip 41, the types of the positive control tissues are 4-100. Can be 4-10, 10-20, 20-30, 30-40, 40-50, 50-60, 60-70, 70-80, 80-90, 90-100.
Further, in the positive control tissue chip 41, the positive control tissue is a human tissue. The gene is homologous with the tissue to be detected, endogenous and high in accuracy.
For example, lung tissue, brain tissue, tonsil tissue, thyroid tissue, thymoma tissue, lung tissue, HER2(3+) breast cancer tissue, liver tissue, myometrium tissue, colon tissue, prostate tissue, brain tissue, stomach tissue, ovary tissue, lymph node tissue, etc. infected with a virus may be mentioned.
Alternatively, the positive control tissue comprises COVID-19 tissue positive for the novel coronavirus antigen. Can be used for the comparison of new coronavirus in a target object to be detected.
In the positive tissue chip 41, each positive control tissue contains 10 to 100 kinds of cells. Can be 10-20, 20-30, 30-40, 40-50, 50-60, 60-70, 70-80, 80-90, 90-100.
The positive tissue chip provided by the invention covers all common tissues and cells of a human body.
The positive control tissue can be flexibly selected according to the needs, and meets various detection requirements.
In one embodiment, the slide substrate 1 of the label area 5 is covered with a chemical resistant coating 51.
Optionally, the chemical-corrosion-resistant coating 51 is composed of a chemical-corrosion-resistant material. The chemical corrosion resistant material is the prior art. For example, the coating described in CN101824266B may be used, and other coatings used in the prior art may also be used.
The label area may be white to facilitate writing.
The label area can be used for pasting the two-dimensional code.
The glass slide substrate of the label area can be a frosted glass slide substrate, so that writing and pasting are facilitated.
Optionally, the area of the label region 5 occupies 25% -32% of the area of the slide. Specifically, 25% -28%, 28% -30% and 30% -32%. Is convenient for writing or pasting, and does not occupy too much area of the glass slide.
Optionally, the area of the control zone 4 is 18-25% of the area of the slide. Specifically, 18% -21%, 21% -23% and 23% -25%. Ensuring enough space for placing the positive tissue chip.
Optionally, in the control zone 4, the length of the positive tissue chip 41 accounts for 16% -17% of the length of the slide.
The positive tissue chips 41 are equal in length and width. Is convenient for manufacturing.
For example, the slide length is 75mm 25mm, the label region is 23mm 25mm, the control region is 18 mm 25mm, the positive tissue chip is 12mm, and the detection region is 34mm 25 mm.
The aforementioned slide glass for immunohistochemical detection can be used for immunohistochemical detection. Including applications in the fields of pathology, immunity, and molecular biology.
Optionally, the method can be used for comparison of the new coronavirus COVID-19 in the target to be detected.
The preparation method of the slide glass for immunohistochemical detection at least comprises the following steps:
1) covering at least part of the surface of the glass slide substrate with an adhesive substance;
2) plating the positive tissue chip on the control area;
3) the surface of the positive tissue chip is covered with anti-degradation substances.
The adherent substance may be applied to the surface of the slide substrate by smearing or dipping.
The surface of the positive tissue chip can be covered with the degradation-preventing substance by smearing or soaking. The method further comprises the steps of:
and coating a chemical corrosion resistant coating on the surface of the glass slide substrate in the label area.
The positive tissue chip can be manufactured by adopting a tissue chip manufacturing method, for example:
the positive control tissue is prepared into a tissue wax block, tens to hundreds of small cylindrical tissues (tissue cores) are collected from a plurality of tissue wax blocks (called donor wax blocks) by a method of fine needle punching by a tissue chip making machine, and are arranged in order into another blank wax block (called receptor wax blocks) and then the tissue chip wax blocks are sliced (thickness is 5um) by a microtomy technology to obtain the positive tissue chip.
For example, methods employing the following steps may also be performed using laboratory instruments, such as an instrumented Lycra bake machine (L EICAHI1220), a Lycra dewaxing all-in-one (L EICA ST5020), and a Dako staining machine (Autostainer L ink 48):
1. dewaxing to water
① placing the tissue slices in a 65 deg.C baking machine for about one hour, ② soaking the slices in xylene for 10min, taking out, and soaking in xylene for 10 min;
③ soaking slices in anhydrous ethanol for 3-5 min, and soaking in 90%, 80% and 70% ethanol for 3-5 min.
2. Antigen retrieval (CBS PH 6.0)
① rinsing the alcohol on the surface of the slice with distilled water for 2-3 times, and leaving no alcohol as much as possible;
② placing the slices into CBS buffer solution preheated in electric cooker (boiling water bath), boiling in water bath for 20min, and switching the electric cooker to heat preservation stage for 10 min;
③ taking out the box and cooling naturally to room temperature.
3. Dropping hydrogen peroxide
① soaking the tissue in PBS buffer solution with pH of 7.2-7.4 for 3 times, each time for 3-5 min;
② taking out the section from the slide rack, manually throwing away PBS buffer solution on the slide, and sucking the buffer solution on the periphery of the tissue by using absorbent paper;
③ placing the slices on a wet box, and dripping 1-2 drops of 3% H2O2And incubating the solution at room temperature for 10-15 min.
4. Drop-wise addition of primary antibody
① placing the slices on a slide rack, soaking the tissue in PBS buffer solution with pH of 7.2-7.4 for 3 times, each time for 3-5 min;
② taking out the section from the slide rack, manually throwing away PBS buffer solution on the slide, and sucking the buffer solution on the periphery of the tissue by using absorbent paper;
③ sections were placed in a wet box, primary antibody was added dropwise, and incubated at 37 ℃ for 60 min.
5. Dropwise adding a second antibody
① placing the slices on a slide rack, soaking the tissue in PBS buffer solution with pH of 7.2-7.4 for 3 times, each time for 3-5 min;
② taking out the section from the slide rack, manually throwing away PBS buffer solution on the slide, and sucking the buffer solution on the periphery of the tissue by using absorbent paper;
③ sections were placed in a wet box, secondary antibody was added dropwise and incubated at room temperature for 30 min.
6. DAB color development
① placing the slices on a slide rack, soaking the tissue in PBS buffer solution with pH of 7.2-7.4 for 3 times, each time for 3-5 min;
② preparing DAB color development liquid, namely preparing the liquid a, the liquid b, the liquid c and distilled water according to the volume ratio of 1: 1: 1: 20;
③ taking out the section from the slide rack, manually throwing away PBS buffer solution on the slide, and sucking the buffer solution on the periphery of the tissue by using absorbent paper;
④ placing the slices on white paper, dripping 50-60 ul of DAB, and performing microscopic examination at room temperature to develop color.
7. Hematoxylin counterstain
① placing the slices on a slide rack, and rinsing the DAB solution on the surfaces of the slices for 2-3 times by using tap water;
② soaking the slices in hematoxylin staining jar for 3min, taking out, and washing off residual hematoxylin on the surface with tap water;
③ soaking the slices in 1% hydrochloric acid alcohol for differentiation, quickly taking out, rinsing in tap water for 2-3 times, discarding the rinsing solution, and rinsing the tissue with running water for 10min (returning blue).
8. Dehydration seal
① soaking the slices in 90% ethanol for 5min, taking out, soaking in anhydrous ethanol for 5min, taking out, and taking out;
② the alcohol and resin seal on the surface of the residual tissue are dried by blowing air.
245 cases of immunohistochemical detection are carried out by using the glass slide, 245 cases of positive tissue chips have no slide release and no slide drop, and the slide release prevention rate reaches 100%. As shown in figure 3, the antigen is well preserved without degradation, the location of cell membrane/plasma/nucleus is clear and accurate, and there is no difference compared with the antigen of fresh tissue.
The foregoing embodiments are merely illustrative of the principles and utilities of the present invention and are not intended to limit the invention. Any person skilled in the art can modify or change the above-mentioned embodiments without departing from the spirit and scope of the present invention. Accordingly, it is intended that all equivalent modifications or changes which can be made by those skilled in the art without departing from the spirit and technical spirit of the present invention be covered by the claims of the present invention.
Claims (10)
1. A slide for immunohistochemical detection, comprising at least: a slide substrate (1), said slide substrate (1) being at least partially covered with an adhesion layer (2); the slide glass is equipped with detection zone (3), control area (4) and label area (5), control area (4) are equipped with positive tissue chip (41), positive tissue chip (41) include a plurality of positive control tissue, detection zone (3) and control area (4) cover have adhesive layer (2), positive tissue chip (41) are located adhesive layer's top.
2. The immunohistochemical detection slide according to claim 1, wherein said adhesive layer (2) is composed of an adhesive substance selected from the group consisting of 3-aminopropyl-3-ethoxysilane, poly-L-lysine and gelatin.
3. The immunohistochemical detection slide according to claim 1, wherein said positive tissue chip (41) is covered with a degradation prevention layer (42) on top.
4. A slide for immunohistochemical detection according to claim 3, wherein said degradation prevention layer (42) is composed of a degradation prevention substance selected from the group consisting of paraffin.
5. The slide for immunohistochemical detection according to claim 1, further comprising one or more of the following features:
a. in the positive tissue chip (41), the types of positive control tissues are 4-100;
b. in the positive control tissue chip (41), the positive control tissue is human tissue;
c. in the positive tissue chip (41), each positive control tissue comprises 10-100 cell types;
d. the glass slide substrate (1) of the label area (5) is covered with a chemical corrosion resistant coating;
e. the area of the label area (5) accounts for 25% -32% of the area of the glass slide;
f. the area of the control zone (4) accounts for 18-25% of the area of the glass slide.
6. The immunohistochemical detection slide according to claim 5, further comprising one or more of the following features:
g. in the control area (4), the area of the positive tissue chip (41) accounts for 16-17% of the length of the glass slide;
h. in the positive tissue chip (41), the positive control tissue comprises a COVID-19 tissue positive to the novel coronavirus antigen.
7. The immunohistochemical detection slide according to claim 6, wherein said positive tissue chip (41) has the same length and width.
8. Use of a slide for immunohistochemical detection according to any one of claims 1 to 7 in an immunohistochemical detection.
9. The method for preparing a slide for immunohistochemical detection according to any one of claims 1 to 7, comprising at least the steps of:
1) covering at least part of the surface of the glass slide substrate with an adhesive substance;
2) plating the positive tissue chip on the control area;
3) the surface of the positive tissue chip is covered with anti-degradation substances.
10. The method for preparing a slide for immunohistochemical detection according to claim 9, further comprising the steps of:
and coating a chemical corrosion resistant substance on the surface of the glass slide substrate in the label area.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113092218A (en) * | 2021-03-19 | 2021-07-09 | 北京龙迈达斯科技开发有限公司 | Teaching chip and preparation method thereof |
| CN114002030A (en) * | 2021-10-15 | 2022-02-01 | 北京龙迈达斯科技开发有限公司 | Composite tissue chip and preparation method thereof |
| CN115220210A (en) * | 2022-07-18 | 2022-10-21 | 上海大格生物科技有限公司 | Immunohistochemical quality control glass slide and quality control chip |
-
2020
- 2020-04-21 CN CN202010318770.6A patent/CN111504739A/en active Pending
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113092218A (en) * | 2021-03-19 | 2021-07-09 | 北京龙迈达斯科技开发有限公司 | Teaching chip and preparation method thereof |
| CN114002030A (en) * | 2021-10-15 | 2022-02-01 | 北京龙迈达斯科技开发有限公司 | Composite tissue chip and preparation method thereof |
| CN115220210A (en) * | 2022-07-18 | 2022-10-21 | 上海大格生物科技有限公司 | Immunohistochemical quality control glass slide and quality control chip |
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Application publication date: 20200807 Assignee: Jiangsu Kangke Investment Co.,Ltd. Assignor: SHANGHAI NINTH PEOPLE'S HOSPITAL SHANGHAI JIAOTONG University SCHOOL OF MEDICINE Contract record no.: X2023310000051 Denomination of invention: A glass slide for immunohistochemical detection License type: Common License Record date: 20230414 |