CN113495153A - System reaction liquid, inhibin A quantitative detection kit containing system reaction liquid and using method - Google Patents
System reaction liquid, inhibin A quantitative detection kit containing system reaction liquid and using method Download PDFInfo
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- 108010067471 inhibin A Proteins 0.000 title claims abstract description 62
- 239000012295 chemical reaction liquid Substances 0.000 title claims abstract description 31
- 238000001514 detection method Methods 0.000 title abstract description 23
- 238000000034 method Methods 0.000 title abstract description 13
- 238000006243 chemical reaction Methods 0.000 claims abstract description 18
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- MHAJPDPJQMAIIY-UHFFFAOYSA-N Hydrogen peroxide Chemical group OO MHAJPDPJQMAIIY-UHFFFAOYSA-N 0.000 claims description 26
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- ILXAOQAXSHVHTM-UHFFFAOYSA-M sodium;2-amino-2-(hydroxymethyl)propane-1,3-diol;chloride Chemical compound [Na+].[Cl-].OCC(N)(CO)CO ILXAOQAXSHVHTM-UHFFFAOYSA-M 0.000 claims description 23
- PXIPVTKHYLBLMZ-UHFFFAOYSA-N Sodium azide Chemical compound [Na+].[N-]=[N+]=[N-] PXIPVTKHYLBLMZ-UHFFFAOYSA-N 0.000 claims description 18
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Images
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
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- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/531—Production of immunochemical test materials
- G01N33/532—Production of labelled immunochemicals
- G01N33/535—Production of labelled immunochemicals with enzyme label or co-enzymes, co-factors, enzyme inhibitors or enzyme substrates
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
- G01N33/54306—Solid-phase reaction mechanisms
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
- G01N2333/435—Assays involving biological materials from specific organisms or of a specific nature from animals; from humans
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Abstract
The invention provides a system reaction liquid, a quantitative inhibin A detection kit containing the same and a using method, wherein the system reaction liquid is added with an oxidant and SDS (sodium dodecyl sulfate), and in the process of detecting inhibin A, a sample is not required to be subjected to oxidation treatment independently, so that a reaction mode that the sample reacts while being oxidized is realized; the inhibin A quantitative detection kit provided by the application overcomes the defects of long reaction time, complex operation steps and the like of the existing inhibin A detection method, and has the characteristics of time saving, convenience in operation, high sensitivity and strong specificity.
Description
Technical Field
The invention belongs to the technical field of immunodiagnosis, and particularly relates to a system reaction liquid, a inhibin A quantitative detection kit containing the system reaction liquid and a using method of the system reaction liquid.
Background
Inhibin (inhin, INH) is one of the members of the transforming growth factor-beta (TGF- β) superfamily of cytokines and is a heterodimeric glycoprotein hormone consisting of an alpha-subunit and a beta-subunit linked by disulfide bonds. The B-subunit is divided into two forms of beta A and beta B, and forms inhibin A (INHA) and inhibin B (INHB) through disulfide bond with the alpha-subunit respectively.
Inhibin A is expressed in both male and female bodies, but mainly appears in maternal serum during pregnancy, the concentration of the maternal serum during pregnancy is high, the inhibin A in the maternal serum rises at the early stage of pregnancy, gradually falls after about 10 weeks, the level is relatively stable at 15-20 weeks, and then rises again, and the inhibin A can reach the peak period when the pregnancy is full. Many studies at home and abroad find that inhibin A is a marker which is found in maternal serum in recent years and can be used for prenatal screening of Down syndrome. Mid-pregnancy down screening was performed in the mother with a "triple screening trial" (i.e., three down screens) consisting of AFP, β -HCG, and uE3, whereas inhibin a would be one of the "four screening trials" following three down screens.
At present, enzyme-linked immunosorbent assay (ELISA) detection of inhibin A adopts a double-antibody sandwich method system including biotin-streptavidin, so that on one hand, the reaction time is long, and on the other hand, the operation steps are complex, and the detection is not convenient for clinical development and use.
Disclosure of Invention
In order to solve the problems, the invention provides a system reaction liquid, a inhibin A quantitative detection kit containing the same and a using method thereof, the system reaction liquid provided by the invention is added with an oxidant and SDS, and in the process of detecting inhibin A, a sample is not required to be subjected to oxidation treatment independently, so that a reaction mode of the sample while being oxidized is realized; the inhibin A quantitative detection kit provided by the application overcomes the defects of long reaction time, complex operation steps and the like of the existing inhibin A detection method, and has the characteristics of time saving, convenience in operation, high sensitivity and strong specificity.
The object of the invention is achieved in the following way: a system reaction liquid comprises protective protein, buffer solution, SDS and oxidant.
The system reaction liquid can also comprise a preservative.
In the system reaction liquid, the oxidant is hydrogen peroxide; the buffer solution is Tris-NaCl buffer solution or PBS buffer solution; the protective protein can be at least one of bovine serum albumin and Casein; the preservative is at least one of Proclin300, NaN3 and cason.
The reaction solution of the system contains 8-12g SDS and 172ml of 30% hydrogen peroxide 163-.
The reaction liquid of the system contains 8-12g SDS, 172ml of 30% hydrogen peroxide 163-.
A quantitative inhibin A detection kit comprises the system reaction liquid, and also comprises an envelope plate coated with an anti-inhibin A antibody, a series of gradient calibrator solutions containing inhibin A antigen, and an enzyme conjugate, wherein the enzyme conjugate is an enzyme-labeled monoclonal antibody aiming at inhibin A.
The coated plate is an ELISA plate coated with monoclonal antibodies aiming at the alpha subunit of inhibin A; the enzyme-labeled monoclonal antibody aiming at inhibin A is a horseradish peroxidase-labeled monoclonal antibody aiming at beta A subunit of inhibin A.
The serial gradient calibrator solutions containing inhibin A antigen are serial gradient calibrator solutions containing inhibin A antigen at concentrations of 0pg/ml, 50pg/ml, 100pg/ml, 200pg/ml, 600pg/ml and 1200pg/ml, respectively.
The coating solution used in the coating plate is one of Tris solution with pH7.4 and PBS solution with pH7.4, the concentration of the monoclonal antibody used for the alpha subunit of inhibin A is 2-10ug/ml, and the blocking solution used is Tris-HCl solution with pH7.4 and containing Bovine Serum Albumin (BSA); the buffer solution used in the preparation of the enzyme conjugate is Tris-NaCl buffer solution or PBS buffer solution, the used protective protein is at least one of calf serum, bovine serum albumin and Casein, and the used preservative is at least one of Proclin300, NaN3 and Kathon; the buffer solution used in the preparation of the calibrator is Tris-NaCl buffer solution or PBS buffer solution, the used protective protein is at least one of calf serum, bovine serum albumin and Casein, and the used preservative is at least one of Proclin300, NaN3 and Kathon; the enzyme-labeled anti-inhibin A monoclonal antibody and the enzyme conjugate diluent are mixed according to the volume ratio of 1 (3800-4200) to obtain the enzyme conjugate.
The using method of the inhibin A quantitative detection kit comprises the following steps,
(1) sample adding: taking out a certain amount of numbered coating holes on the coating plate, sequentially adding 50 mul of calibrator and sample to be tested, then respectively adding 50 mul of system reaction solution, and then adding 50 mul of enzyme conjugate;
(2) and (3) incubation: placing the coated plate after sample adding at the ambient temperature of 37 ℃ for 60 minutes;
(3) washing: throwing off liquid in the coating holes, filling the coating holes with diluted washing liquid, standing for 20 seconds, throwing off, repeatedly washing for 5 times in such a way, throwing off the liquid each time, and finally patting on absorbent paper to be dry;
(4) color development: adding 100 mul of substrate solution into each coating hole, and reacting for 20 minutes at 18-23 ℃ in a dark place;
(5) and (4) terminating: the reaction was stopped by adding 50. mu.l of a stop solution to each coated well.
The kit is prepared based on an enzyme-linked immunosorbent assay, is used together with an enzyme-linked immunosorbent assay, and has the advantages of simple and convenient operation and high detection speed. Compared with the inhibin A detection kit on the market, the kit adopts a double-antibody sandwich method for directly labeling an anti-inhibin A antibody by enzyme to realize a one-step reaction mode; the formula of the reaction liquid of the system is optimized, the oxidant and SDS are added, the sample is not required to be oxidized independently, and the reaction mode of oxidizing and reacting the sample is realized.
The hydrogen peroxide in the reaction liquid of the system is used as the main component for oxidizing the sample, and the site containing methionine in inhibin A in the sample is fully exposed, so that the antibody can be better identified and combined, and the obtained detection result is more accurate.
Drawings
FIG. 1 is a reference diagram showing the correlation between the kit of the present application and the existing products.
Detailed Description
A system reaction liquid comprises protective protein, buffer solution, SDS and oxidant.
The system reaction liquid can also comprise a preservative.
In the system reaction liquid, the oxidant is hydrogen peroxide; the buffer solution is Tris-NaCl buffer solution or PBS buffer solution; the protective protein can be at least one of bovine serum albumin and Casein; the preservative is at least one of Proclin300, NaN3 and cason.
The reaction solution of the system contains 8-12g SDS and 172ml of 30% hydrogen peroxide 163-.
The reaction liquid of the system contains 8-12g SDS, 172ml of 30% hydrogen peroxide 163-.
A quantitative inhibin A detection kit comprises the system reaction liquid, and also comprises an envelope plate coated with an anti-inhibin A antibody, a series of gradient calibrator solutions containing inhibin A antigen, and an enzyme conjugate, wherein the enzyme conjugate is an enzyme-labeled monoclonal antibody aiming at inhibin A.
The coated plate is an ELISA plate coated with monoclonal antibodies aiming at the alpha subunit of inhibin A; the enzyme-labeled monoclonal antibody aiming at inhibin A is a horseradish peroxidase-labeled monoclonal antibody aiming at beta A subunit of inhibin A.
The serial gradient calibrator solutions containing inhibin A antigen are serial gradient calibrator solutions containing inhibin A antigen at concentrations of 0pg/ml, 50pg/ml, 100pg/ml, 200pg/ml, 600pg/ml and 1200pg/ml, respectively.
The coating solution used in the coating plate is one of Tris solution with pH7.4 and PBS solution with pH7.4, the concentration of the monoclonal antibody used for the alpha subunit of inhibin A is 2-10ug/ml, and the blocking solution used is Tris-HCl solution with pH7.4 and containing bovine serum albumin; the buffer solution used in the preparation of the enzyme conjugate is Tris-NaCl buffer solution or PBS buffer solution, the used protective protein is at least one of calf serum, bovine serum albumin and Casein, and the used preservative is at least one of Proclin300, NaN3 and Kathon; the buffer solution used in the preparation of the calibrator is Tris-NaCl buffer solution or PBS buffer solution, the used protective protein is at least one of calf serum, bovine serum albumin and Casein, and the used preservative is at least one of Proclin300, NaN3 and Kathon; the enzyme-labeled anti-inhibin A monoclonal antibody and the enzyme conjugate diluent are mixed according to the volume ratio of 1 (3800-4200) to obtain the enzyme conjugate.
The using method of the inhibin A quantitative detection kit comprises the following steps,
(1) sample adding: taking out a certain amount of numbered coating holes on the coating plate, sequentially adding 50 mul of calibrator and sample to be tested, then respectively adding 50 mul of system reaction solution, and then adding 50 mul of enzyme conjugate;
(2) and (3) incubation: placing the coated plate after sample adding at the ambient temperature of 37 ℃ for 60 minutes;
(3) washing: throwing off liquid in the coating holes, filling the coating holes with diluted washing liquid, standing for 20 seconds, throwing off, repeatedly washing for 5 times in such a way, throwing off the liquid each time, and finally patting on absorbent paper to be dry;
(4) color development: adding 100 mul of substrate solution into each coating hole, and reacting for 20 minutes at 18-23 ℃ in a dark place;
(5) and (4) terminating: the reaction was stopped by adding 50. mu.l of a stop solution to each coated well.
The present invention is described in more detail below with reference to specific examples to facilitate understanding by those skilled in the art. If not specifically stated, the materials and the detecting instruments used in the invention are conventional materials in the industry, and are common products sold in the market. The detection method used is a conventional method used in the industry.
Preparation of inhibin A quantitative detection kit
1. Preparation of coated plate
The coating solution is one of Tris with pH7.4, and the antibody is a monoclonal antibody aiming at the alpha subunit of inhibin A, and the concentration is 5 ug/ml. The blocking solution used was 0.05M Tris-HCl pH7.4 containing bovine serum albumin, which was prepared according to a conventional method for preparing a coating antibody. The source manufacturer and model of monoclonal antibody against the alpha subunit of inhibin A are Hangzhou Boyue organism B2.
(1) Preparation of coating liquid: dissolving 6.05g of Tris and 8.5g of NaCl in 900mL of purified water, adjusting the pH value to 7.4 by using 6M HCl, and metering the volume to 1000mL to prepare a coating solution; (2) preparation of a sealing liquid: dissolving 6.05g of Tris and 8.5g of NaCl in 900mL of purified water, adjusting the pH value to 7.4 by using HCl, and metering the volume to 1000mL to obtain a solution A; then, 30g of BSA, 1ml of Proclin300 and 100g of sucrose were added to a portion of solution A, and then the volume was increased to 1000ml by adding solution A to prepare a blocking solution.
Tris, NaCl, BSA, purified water, sucrose, Proclin300 were analytically pure, respectively.
2. Preparation of enzyme conjugates
The buffer solution of the enzyme conjugate adopts Tris-NaCl buffer solution, the used protective protein can be bovine serum albumin, and the used preservative is Proclin 300.
(1) Preparation of Tris-NaCl buffer: dissolving 6.05g of Tris and 8.5g of NaCl in 900mL of purified water, adjusting the pH value to 7.4 by using 6M HCl, and metering the volume to 1000 mL;
(2) firstly, taking a part of Tris-NaCl buffer solution, adding 30g of BSA and 1ml of Proclin300 into the Tris-NaCl buffer solution, and then carrying out constant volume to 1000ml by adding the Tris-NaCl buffer solution to prepare an enzyme conjugate diluent;
(3) and mixing the HRP-labeled anti-inhibin A monoclonal antibody with the enzyme conjugate diluent according to the volume ratio of 1:4000 to obtain the enzyme conjugate.
Tris, NaCl, BSA, purified water, Proclin300 were analytically pure, respectively. The source and type of the anti-inhibin A monoclonal antibody are Hangzhou Boyue organism and B13 respectively.
Adding an HRP-labeled anti-inhibin A monoclonal antibody into the enzyme conjugate diluent according to the ratio of 1:4000 to obtain the enzyme conjugate.
3. Preparation of the systematic reaction solution
The buffer solution of the reaction solution of the system adopts Tris-NaCl buffer solution, the protective protein is bovine serum albumin, and the used preservative adopts Proclin 300.
(1) Preparation of Tris-NaCl buffer: dissolving 6.05g of Tris and 8.5g of NaCl in 700mL of purified water, and adjusting the pH value to 7.4 by using 6M HCl;
(2) then adding 167mL of 30% hydrogen peroxide, and then fixing the volume to 1000 mL;
(3) firstly, taking a part of Tris-NaCl buffer solution, adding 10g of BSA, 1ml of Proclin300 and 10g of sodium dodecyl sulfate into the Tris-NaCl buffer solution, and then carrying out constant volume to 1000ml by adding the Tris-NaCl buffer solution to prepare the reaction solution of the system.
Tris, NaCl, BSA, 30% hydrogen peroxide, purified water, sodium dodecyl sulfate and Proclin300 are analytically pure respectively.
4. Preparation of calibrator
(1) Preparation of Tris-NaCl buffer: dissolving 6.05g of Tris and 8.5g of NaCl in 700mL of purified water, adjusting the pH value to 7.4 by using 6M HCl, and metering to 1000 mL;
(2) firstly, taking a part of Tris-NaCl buffer solution, adding 30g of BSA, 1ml of Tween-20, 5g of gentamicin and 1ml of Proclin300 into the Tris-NaCl buffer solution, and then fixing the volume to 1000ml by adding the Tris-NaCl buffer solution to prepare the calibrator diluent.
Tris, NaCl, BSA, gentamicin, purified water, inhibin A, Proclin300 and Tween-20 are analytically pure respectively.
The inhibin A is diluted by the calibrator diluent to prepare six concentrations of inhibin A calibrator, namely 0pg/ml, 50pg/ml, 100pg/ml, 200pg/ml, 600pg/ml and 1200 pg/ml. The inhibin A antigen is from Hangzhou Boyue organism.
5. Other general reagent components: the substrate, stop solution and washing solution are purchased from Zhengzhou Danuo organisms.
6. Procedure of the test
1. Experimental procedure
(1) Sample adding: taking out a certain amount of numbered coating holes on the coating plate: and sequentially adding 50 mul of calibrator and sample to be detected by using a micropipette, respectively adding 50 mul of system reaction solution, and then adding 50 mul of enzyme conjugate.
(2) And (3) incubation: the coated plate after loading was placed at 37 ℃ for 60 minutes.
(3) Washing: throwing off liquid in the coating holes, filling the coating holes with diluted washing liquid, standing for 20 seconds, throwing off, washing for 5 times repeatedly, throwing off liquid every time, and finally patting on absorbent paper.
(4) Color development: add 100. mu.l of substrate solution to each coated well, and react for 20 minutes at room temperature 18-23 ℃ in the dark.
(5) And (4) terminating: the reaction was stopped by adding 50. mu.l of a stop solution to each coated well.
2. Calculation of results
(1) Detection by a microplate reader: the microplate reader wavelength is 450nm, and the reference wavelength is 630 nm. The OD of each well was measured within 10 minutes after the termination of the reaction.
(2) And (3) calculating: the kit recommends a point-to-point regression fitting mode, namely, a point-to-point standard curve is established by taking the concentration value of a series of calibration products as an abscissa (X axis) and the OD value of a standard product as an ordinate (Y axis), so that the content of the inhibin A in a sample to be detected is calculated according to the curve.
Exemplary data were calculated as in table 1 below.
TABLE 1
3. Clinical verification of the kit of the invention
In order to verify the consistency of the results of inhibin A in the kit of the invention and anshlabs of a comparative manufacturer, 103 serum samples are collected for parallel detection, the detection results are analyzed for clinical relevance, the results are shown in the following table 2, and a reference graph for the relevance is prepared according to the data in the table and is shown in fig. 1.
TABLE 2
The comparison results shown in fig. 1 indicate that the currently relevant R2 of the kit of the invention and the comparative kit is 0.9786, which proves that the kit of the invention and the comparative kit have better correlation.
The kit adopts a one-step reaction, the horseradish peroxidase is directly coupled on the anti-inhibin A monoclonal antibody, and the system reaction liquid added with the hydrogen peroxide and the enzyme conjugate are simultaneously added, so that the oxidation of a sample site and the combination of the antibody are simultaneously carried out, the reaction time is reduced, the operation steps are simplified, the more convenient and faster operation method is realized, and the kit is more flexible and convenient for clinical operation and use.
The foregoing is only a preferred embodiment of the present invention, and it should be noted that, for those skilled in the art, various changes and modifications can be made without departing from the overall concept of the present invention, and these should also be considered as the protection scope of the present invention.
Claims (10)
1. A system reaction solution, which is characterized in that: comprises protective protein, buffer solution, SDS and oxidant.
2. The system reaction liquid according to claim 1, characterized in that: preservatives may also be included.
3. The system reaction liquid according to claim 2, characterized in that: the oxidant is hydrogen peroxide; the buffer solution is Tris-NaCl buffer solution or PBS buffer solution; the protective protein can be at least one of bovine serum albumin and Casein; the preservative is at least one of Proclin300, NaN3 and cason.
4. The system reaction liquid according to claim 1, characterized in that: each liter contains 8-12g SDS, 30% hydrogen peroxide 163-172 ml.
5. The system reaction liquid according to claim 1, characterized in that: each liter contains 8-12g SDS, 30% hydrogen peroxide 163-172ml, bovine serum albumin 8-12g, Proclin 3000.8-1.3 ml.
6. A inhibin A quantitative determination kit is characterized in that: the system reaction liquid comprises the system reaction liquid of any one of claims 1 to 5, and further comprises a coating plate coated with an anti-inhibin A antibody, a series of gradient calibrator solutions containing inhibin A antigen, and an enzyme conjugate, wherein the enzyme conjugate is an enzyme-labeled monoclonal antibody aiming at inhibin A.
7. The kit for quantitatively detecting inhibin A according to claim 6, characterized in that: the coated plate is an ELISA plate coated with monoclonal antibodies aiming at the alpha subunit of inhibin A; the enzyme-labeled monoclonal antibody aiming at inhibin A is a horseradish peroxidase-labeled monoclonal antibody aiming at beta A subunit of inhibin A.
8. The kit for the quantitative determination of inhibin A according to claim 6 or 7, wherein: the serial gradient calibrator solutions containing inhibin A antigen are serial gradient calibrator solutions containing inhibin A antigen at concentrations of 0pg/ml, 50pg/ml, 100pg/ml, 200pg/ml, 600pg/ml and 1200pg/ml, respectively.
9. The kit for the quantitative determination of inhibin A according to claim 6 or 7, wherein: the coating solution used in the coating plate is one of Tris solution with pH7.4 and PBS solution with pH7.4, the concentration of the monoclonal antibody used for the alpha subunit of inhibin A is 2-10ug/ml, and the blocking solution used is Tris-HCl solution with pH7.4 and containing Bovine Serum Albumin (BSA); the buffer solution used in the preparation of the enzyme conjugate is Tris-NaCl buffer solution or PBS buffer solution, the used protective protein is at least one of calf serum, bovine serum albumin and Casein, and the used preservative is at least one of Proclin300, NaN3 and Kathon; the buffer solution used in the preparation of the calibrator is Tris-NaCl buffer solution or PBS buffer solution, the used protective protein is at least one of calf serum, bovine serum albumin and Casein, and the used preservative is at least one of Proclin300, NaN3 and Kathon; the enzyme-labeled anti-inhibin A monoclonal antibody and the enzyme conjugate diluent are mixed according to the volume ratio of 1 (3800-4200) to obtain the enzyme conjugate.
10. The use of the kit for the quantitative determination of inhibin A according to any one of claims 6-9, characterized in that: comprises the following steps (1),
sample adding: taking out a certain amount of numbered coating holes on the coating plate, sequentially adding 50 mul of calibration material and a sample to be detected, then respectively adding 50 mul of system reaction liquid, and then adding 50 mul of enzyme conjugate;
(2) and (3) incubation: placing the coated plate after sample adding at the ambient temperature of 37 ℃ for 60 minutes;
(3) washing: throwing off liquid in the coating holes, filling the coating holes with diluted washing liquid, standing for 20 seconds, throwing off, repeatedly washing for 5 times in such a way, throwing off the liquid each time, and finally patting on absorbent paper to be dry;
(4) color development: adding 100 mul of substrate solution into each coating hole, and carrying out a light-shielding reaction for 20 minutes at 18-23 ℃;
(5) and (4) terminating: adding 50 mu l of stop solution into each coating hole to stop the reaction.
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| US5952182A (en) * | 1994-05-24 | 1999-09-14 | Oxford Brookes University | Method of genetic testing |
| US20090317921A1 (en) * | 2008-06-11 | 2009-12-24 | Oxford Brookes University | Antibody to inhibin/ activin beta-b subunit |
| CN106199012A (en) * | 2016-06-30 | 2016-12-07 | 深圳市亚辉龙生物科技股份有限公司 | Inhibin B chemiluminescence immune detection reagent kit and preparation method thereof |
| CN109061198A (en) * | 2018-08-17 | 2018-12-21 | 迪瑞医疗科技股份有限公司 | inhibin A detection kit and preparation method thereof |
| CN109444432A (en) * | 2018-12-15 | 2019-03-08 | 郑州安图生物工程股份有限公司 | A kind of kit detecting inhibin A |
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| US5952182A (en) * | 1994-05-24 | 1999-09-14 | Oxford Brookes University | Method of genetic testing |
| US20090317921A1 (en) * | 2008-06-11 | 2009-12-24 | Oxford Brookes University | Antibody to inhibin/ activin beta-b subunit |
| CN106199012A (en) * | 2016-06-30 | 2016-12-07 | 深圳市亚辉龙生物科技股份有限公司 | Inhibin B chemiluminescence immune detection reagent kit and preparation method thereof |
| CN109061198A (en) * | 2018-08-17 | 2018-12-21 | 迪瑞医疗科技股份有限公司 | inhibin A detection kit and preparation method thereof |
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