CN111505297B - Application of endosulfan artificial antigen in enzyme linked immunosorbent assay kit - Google Patents
Application of endosulfan artificial antigen in enzyme linked immunosorbent assay kit Download PDFInfo
- Publication number
- CN111505297B CN111505297B CN202010400219.6A CN202010400219A CN111505297B CN 111505297 B CN111505297 B CN 111505297B CN 202010400219 A CN202010400219 A CN 202010400219A CN 111505297 B CN111505297 B CN 111505297B
- Authority
- CN
- China
- Prior art keywords
- endosulfan
- solution
- enzyme
- immunosorbent assay
- linked immunosorbent
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
Images
Classifications
-
- 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
- G01N33/577—Immunoassay; Biospecific binding assay; Materials therefor involving monoclonal antibodies binding reaction mechanisms characterised by the use of monoclonal antibodies; monoclonal antibodies per se are classified with their corresponding antigens
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/76—Albumins
- C07K14/765—Serum albumin, e.g. HSA
-
- 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
- G01N33/5308—Immunoassay; Biospecific binding assay; Materials therefor for analytes not provided for elsewhere, e.g. nucleic acids, uric acid, worms, mites
-
- 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
- G01N33/58—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving labelled substances
- G01N33/581—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving labelled substances with enzyme label (including co-enzymes, co-factors, enzyme inhibitors or substrates)
Abstract
The invention provides an enzyme linked immunosorbent assay kit for detecting endosulfan, which comprises: the enzyme-linked immunosorbent assay kit comprises an ELISA plate coated with a coating antigen, a endosulfan standard solution, endosulfan antibodies, an enzyme conjugate concentrated solution, an enzyme conjugate diluent, a substrate color developing solution, a stop solution and a washing solution, wherein the coating antigen is endosulfan coupled antigen, the enzyme conjugate is an enzyme-labeled endosulfan antibody, and the endosulfan antibody is obtained by immunizing an animal with immunogen. The invention also discloses a method for detecting endosulfan by using the enzyme linked immunosorbent assay kit, which comprises the following steps: firstly, preprocessing a sample, then detecting by using a kit, and finally analyzing a detection result. The enzyme linked immunosorbent assay kit provided by the invention can be used for detecting the content of endosulfan in vegetable and fruit samples, is simple and convenient to operate, low in cost, high in sensitivity, capable of being monitored on site and suitable for screening a large number of samples.
Description
Technical Field
The invention relates to an application technology of a endosulfan artificial antigen in an enzyme-linked immunoassay kit, in particular to an endosulfan artificial antigen molecular structure and an enzyme-linked immunoassay kit for detecting endosulfan, which can qualitatively and quantitatively detect the residual quantity of endosulfan medicines in vegetables and fruits.
Background
Endosulfan is an artificially synthesized organochlorine compound and is widely used as a pesticide. The pesticide has the advantages of wide pesticidal range, low cost, low toxicity to bees, etc., and can be widely applied to various crops. China began to produce endosulfan in the last 90 s, and by 2011, 47 endosulfan products which are registered and used relate to 43 families of medicine enterprises, which are called the second major producing country of the medicine. The medicine has long shelf life, biological enrichment effect, high harm to fish and shellfish, and cumulative toxicity to mammal. There have been a number of reports in the literature on the detection of endosulfan residues in crops such as vegetables and fruits.
The method for researching endosulfan at home and abroad mainly adopts solid-phase extraction and gas chromatography detection. Compared with the two methods, the immunochemical detection method has the advantages of rapidness, specificity, sensitivity, accuracy, batch monitoring, simple sample treatment, automatic operation and the like for the detection of the medicine. The invention discloses a method for preparing artificial antigen of endosulfan, which is applied to an enzyme-linked immunosorbent assay kit, has the advantages of short time, simple operation and lower cost, and is suitable for detecting samples in bulk by a basic unit.
Disclosure of Invention
The invention aims to provide an enzyme linked immunosorbent assay kit capable of detecting the residual quantity of endosulfan in vegetables and fruits, and provides a qualitative and quantitative detection method which is efficient, accurate, simple and convenient and is suitable for screening large-batch samples.
The kit of the invention comprises: the enzyme-linked immunosorbent assay kit comprises an ELISA plate coated with a coating antigen, a endosulfan standard solution, endosulfan antibodies, an enzyme conjugate concentrated solution, an enzyme conjugate diluent, a substrate color developing solution, a stop solution and a washing solution, wherein the coating antigen is endosulfan coupled antigen, and the enzyme conjugate is enzyme-labeled endosulfan antibodies.
The endosulfan specific antibody is prepared by taking an endosulfan artificial antigen as an immunogen, and can be an endosulfan monoclonal antibody or an endosulfan polyclonal antibody, wherein the endosulfan monoclonal antibody is preferred.
The labeled enzyme of the enzyme conjugate is horseradish peroxidase or alkaline phosphatase extracted from bacteria, wherein horseradish peroxidase is preferred; the enzyme conjugate is obtained by coupling enzyme and endosulfan antibody.
In order to facilitate field monitoring and large-scale sample screening, the kit further comprises a endosulfan standard solution, a substrate developing solution, a stop solution and a washing solution.
6 bottles of endosulfan standard substance solution are respectively 0 mug/L, 1 mug/L, 3 mug/L, 9 mug/L, 27 mug/L and 81 mug/L in concentration.
When the labeled enzyme is horseradish peroxidase, the substrate color development solution consists of a substrate solution A and a substrate solution B, wherein A is hydrogen peroxide or carbamide peroxide, the B is o-phenylenediamine or tetramethyl benzidine, and the stop solution is 1 to 2mol/L sulfuric acid solution or hydrochloric acid buffer solution; when the marker enzyme is bacteria to extract alkaline phosphatase, the substrate color developing solution is a para-nitrophosphate buffer solution, and the stop solution is 1 to 2mol/L sodium hydroxide solution.
The washing liquid preferably has a pH value of 7.4, and contains 0.5% -1.0% of Tween-20, 0.01% -0.03% of sodium azide preservative and 0.1-0.3 mol/L of phosphate buffer solution, wherein the percentage is weight volume percentage.
The enzyme label plate is characterized in that a coating buffer solution used in the preparation process of the enzyme label plate is a carbonate buffer solution with the pH value of 9.6 and 0.05mol/L, a confining solution is a phosphate buffer solution with the pH value of 7.1 to 7.5 and contains 1 to 3 percent of casein and 0.1 to 0.3mol/L, and the percentage is weight volume percentage.
The preparation process of the ELISA plate comprises the following steps: diluting the coating source into 20 mu g/mL by using a coating buffer solution, adding 100 mu l of the coating source into each hole, incubating for 2 hours at 25 ℃ in the dark or overnight at 4 ℃, pouring out liquid in the holes, washing for 2 times by using a washing solution for 30s each time, patting to dry, then adding 150 to 200 mu l of sealing liquid into each hole, incubating for 1 to 2h in the dark at 25 ℃, pouring out liquid in the holes, patting to dry, drying, and then sealing and storing in vacuum by using an aluminum film.
The detection principle of the invention is as follows:
the kit adopts a direct competition ELISA method, pre-coats a coupling antigen on an enzyme label plate micropore strip, the residual endosulfan in a sample and the coupling antigen pre-coated on the enzyme label plate micropore strip compete for an enzyme conjugate resisting endosulfan, a TMB substrate is used for developing color, the absorbance value of the sample is in negative correlation with the content of the residual endosulfan in the sample, the absorbance value is compared with a standard curve, and the absorbance value is multiplied by the corresponding dilution multiple to obtain the residual amount of the endosulfan in the sample.
The invention also provides a method for detecting endosulfan by using the enzyme linked immunosorbent assay kit, which comprises the following steps:
(1) Pretreating a sample;
(2) Detecting with the kit;
(3) And analyzing the detection result.
The enzyme linked immunosorbent assay kit for detecting the endosulfan mainly adopts an ELISA method to qualitatively or quantitatively detect the content of the endosulfan in a sample; the pretreatment requirement on the sample is low, the pretreatment process of the sample is simple, and a large amount of samples can be detected quickly; the main reagent is provided in the form of working solution, and the detection method is convenient and easy to implement and has the characteristics of high specificity, high sensitivity, high precision, high accuracy and the like. The enzyme linked immunosorbent assay kit has the advantages of simple structure, convenient use, low price, convenient carrying, high efficiency, accuracy, simplicity and convenience of a detection method, and suitability for qualitative and quantitative screening of large-batch samples.
Drawings
FIG. 1: molecular structural formula of endosulfan hapten
Detailed Description
The invention is further illustrated below with reference to specific examples. It should be understood that these examples are for illustrative purposes only and are not intended to limit the scope of the present invention.
EXAMPLE 1 preparation of kit Components
1. Preparation of endosulfan hapten
Taking 3.6g of endosulfan, adding 100ml of 1, 4-dioxane for dissolving, fully stirring, then adding 3.0g of molecular sieve, adding 1.02g of succinic semialdehyde, fully stirring, introducing hydrochloric acid gas, and stirring for 5 hours at room temperature in a dark place; stopping the reaction, adding 0.5mol/L NaOH solution, fully shaking 200ml, adding 200ml of ethyl acetate, fully shaking, standing, separating a water phase, washing an organic phase with 80ml of water, standing after shaking, separating the water phase, drying the organic phase with anhydrous sodium sulfate, drying to dryness, loading on a silica gel column, and eluting and purifying dichloromethane/methanol (v/v, 10/1) to obtain 2.1g of the succinyl acetal endosulfan hapten product with the yield of 45.75%.
2. Preparation of antigens
Immunogen preparation-endosulfan hapten is coupled with Bovine Serum Albumin (BSA) to obtain the immunogen.
Dissolving Succinal acetal endosulfan 16mg in DMF 1ml, adding NHS18.7mg and DDC33mg, reacting at room temperature for 3 hr to obtain hapten solution A; taking 50mg of Bovine Serum Albumin (BSA), adding 0.05M PB buffer solution for dissolving to obtain solution B, dripping A solution into the solution B, reacting at 4 ℃ for 12h, dialyzing and purifying by 0.02M PBS for 3 days, and changing the solution three times every day to obtain the endosulfan-BSA conjugate which is the immunogen and storing at-20 ℃ for later use.
Preparation of coating antigen-coupling of endosulfan hapten and Ovalbumin (OVA) to obtain immunogen.
Dissolving Succinal acetal endosulfan 10mg in DMSO 1ml, adding NHS6.7mg and EDC16mg, reacting at room temperature for 3 hr to obtain hapten solution A; dissolving egg serum albumin (OVA) 50mg in 0.05M PB buffer solution to obtain solution B, adding solution A into solution B, reacting at 4 deg.C for 12h, dialyzing and purifying with 0.02M PBS for 3 days, and changing solution three times per day to obtain endosulfan-OVA conjugate, which is coating antigen, and storing at-20 deg.C.
3. Preparation of endosulfan monoclonal antibody
Animal immunization: injecting the immunogen obtained in the above steps into Balb/c mice, wherein the immunization dose is 150 mug/mouse, so that antiserum is generated.
Cell fusion and cloning: after the measurement result of the mouse serum is higher, spleen cells are taken and fused with SP2/0 myeloma cells according to the proportion of 8 (quantitative ratio), cell supernatants are measured by adopting indirect competition ELISA, and positive holes are screened. Cloning the positive hole by using a limiting dilution method until obtaining a hybridoma cell strain secreting the endosulfan monoclonal antibody.
Freezing and recovering cells: preparation of monoclonal hybridoma cell line into 1 × 10 frozen stock solution 6 Cell suspension per mL, stored for long periods in liquid nitrogen. Taking out the frozen tube during recovery, immediately putting the tube into a water bath at 37 ℃ for quick melting, centrifuging to remove frozen stock solution, and transferring the tube into a culture bottle for culture.
Production and purification of monoclonal antibodies: injecting sterilized paraffin oil 0.5 mL/mouse into Balb/c mouse abdominal cavity, injecting stable monoclonal hybridoma cell strain 5X 10 into the abdominal cavity after 7 days 5 Ascites were collected 7 days later. Purifying ascites by octanoic acid-saturated ammonium sulfate method, and storing at-20 deg.C.
4. Preparation of enzyme conjugates
Taking a goat as an immune animal, and taking a endosulfan monoclonal antibody as an immunogen to immunize a pathogen-free goat, so as to obtain the endosulfan antibody. And (3) coupling the endosulfan antibody with horseradish peroxidase (HRP) to obtain an enzyme conjugate.
5. Preparation of ELISA plate
Diluting the coating source to 20 mu g/mL by using a coating buffer solution, adding 100 mu l of the coating source into each hole, incubating for 2h at 25 ℃ in a dark place, pouring off liquid in the holes, washing for 2 times by using a washing solution for 30s each time, patting to dry, then adding 200 mu l of a sealing solution into each hole, incubating for 2h at 25 ℃ in a dark place, pouring off liquid in the holes, patting to dry, and performing vacuum sealing and storage by using an aluminum film after drying.
EXAMPLE 2 construction of enzyme-linked immunoassay kit for detecting endosulfan
An enzyme linked immunosorbent assay kit for detecting endosulfan is constructed, and comprises the following components:
(1) An ELISA plate coated with endosulfan coupling antigen;
(2) 6 bottles of endosulfan standard substance solution with the concentration of 0 mug/L, 1 mug/L, 3 mug/L, 9 mug/L, 27 mug/L and 81 mug/L respectively;
(3) Using sulfur-lead antibody marked by horseradish peroxidase;
(4) The substrate color development liquid consists of a liquid A and a liquid B, wherein the liquid A is carbamide peroxide, and the liquid B is tetramethyl benzidine;
(5) The stop solution is 2mol/L sulfuric acid;
(6) The washing liquid has a pH value of 7.4, and contains 0.5-1.0% of Tween-20, 0.01-0.03% of sodium azide preservative and 0.1-0.3 mol/L of phosphate buffer solution, wherein the percentage is weight volume percentage;
example 3 detection of endosulfan in vegetables and fruits
1. Detection with a kit
And (3) numbering the corresponding micropores of the samples and the standard products in sequence, making 2 holes of each sample and each standard product in parallel, and recording the positions of the standard holes and the sample holes. Adding 50ml of standard substance/sample into the corresponding micropores, then adding 50ml of enzyme conjugate working solution into the micropores, and reacting for 30min at 25 ℃ in a dark environment. And (5) spin-drying the liquid in the holes, washing for 4-5 times, and patting dry. Adding 50 ml/hole of the substrate solution A, adding 50 ml/hole of the substrate solution B, mixing, and reacting at 25 deg.C in a dark environment for 15min. Adding 50ml of stop solution per well, mixing uniformly, setting an enzyme-labeling instrument at 450nm, and measuring the OD value of each well.
2. Analysis of detection results
The percent absorbance of the standard or sample is equal to the average of the absorbance values of the standard or sample (double well) divided by the average of the absorbance values of the first standard (0 standard) and multiplied by 100% to obtain the percent absorbance value of the standard or sample. And drawing a standard curve graph by taking the percent absorbance of the standard substance as a vertical coordinate and taking the logarithm of the concentration (mu g/L) of the endosulfan standard substance as a horizontal coordinate. And substituting the percent absorbance of the sample into the standard curve, reading out the concentration corresponding to the sample from the standard curve, and multiplying the corresponding dilution times to obtain the actual concentration of the endosulfan in the sample.
EXAMPLE 4 determination of endosulfan technical parameters
1. Sensitivity and detection limits of the kit
Determining the sensitivity of the kit according to a conventional method, wherein the range of a standard curve is 1 to 81 mug/L and IC 50 The floating range (50% inhibition concentration) is 4.5 to 7.5 mu g/L; the 20 samples are tested, the concentration corresponding to each percent absorbance value is found out from the standard curve, the detection limit is represented by adding 3 times of standard deviation to the average value of the 20 sample concentration, and the result shows that the detection limit of the method for endosulfan in vegetables and fruits is respectively 50 mug/kg and 100 mug/kg.
2. Sample precision and accuracy testing
The recovery rate is used as an accuracy evaluation index, and the relative standard deviation (RSD%) of the detection result of a sample with a certain concentration is repeatedly measured and used as a precision evaluation index. The calculation formula is as follows: recovery (%) = actual measurement value/theoretical value × 100%, where the theoretical value is the added concentration of the sample; relative standard deviation RSD% = SD/X × 100%, where SD is the standard deviation and X is the average of the measured data.
Adding and recovering the vegetable samples according to the endosulfan with the concentration of 50 mug/kg and 100 mug/kg, adding and recovering the fruit samples according to the endosulfan with the concentration of 100 mug/kg and 200 mug/kg, performing adding and recovering determination on the fruit samples, performing determination on each sample by 4 parallels, using three different reagents, and calculating the average recovery rate and the precision result of the samples, wherein the results are shown in the following table.
TABLE 1 vegetable and fruit sample precision and accuracy tests
Adding and recovering measurement is carried out on the vegetable sample according to endosulfan with the concentration of 50 mug/kg and 100 mug/kg, adding and recovering measurement is carried out on the fruit sample according to endosulfan with the concentration of 100 mug/kg and 200 mug/kg, and the average recovery rates are 81.2% -92.3% and 74.8% -87.9% respectively; the relative standard deviation in each batch and between batches is less than 15 percent.
3. Test of stability of kit
The storage condition of the kit is 2-8 ℃, and the maximum absorbance value (zero standard), the 50% inhibition concentration and the actual measurement value of the added endosulfan of the kit are all within the normal range through measurement for 12 months. Considering that abnormal storage conditions occur in the transportation and use processes, the kit is placed for 7 days under the storage condition of 37 ℃ for accelerated aging experiments, and the result shows that all indexes of the kit completely meet the requirements. And in consideration of the occurrence of the freezing condition of the kit, the kit is frozen for 7 days in a refrigerator at the temperature of-20 ℃, and the determination result also shows that all indexes of the kit are completely normal. From the above results, it was found that the kit can be stored at 2 to 8 ℃ for at least 12 months.
Claims (3)
1. An enzyme linked immunosorbent assay kit for detecting endosulfan is characterized by comprising: the enzyme-linked immunosorbent assay kit comprises an ELISA plate coated with a coating antigen, a endosulfan standard solution, endosulfan antibodies, an enzyme conjugate concentrated solution, an enzyme conjugate diluent, a substrate color developing solution, a stop solution and a washing solution, wherein the coating antigen is an endosulfan conjugate antigen, the enzyme conjugate is an enzyme-labeled endosulfan antibody, the endosulfan antibody is obtained by immunizing animals with immunogen, the endosulfan conjugate antigen is obtained by coupling endosulfan hapten and carrier protein, the endosulfan hapten is obtained by carrying out a series of chemical reactions on endosulfan, 1, 4-dioxane and succinsemialdehyde, and the molecular structural formula of the endosulfan hapten is as follows:
2. the kit of claim 1, characterized in that the immunogen is prepared as follows:
dissolving Succinal acetal endosulfan 16mg in DMF 1ml, adding NHS18.7mg and DDC33mg, and reacting at room temperature for 3 hr to obtain hapten solution A; taking 50mg of bovine serum albumin BSA, adding 0.05M PB buffer solution for dissolving to obtain solution B, dripping the solution A into the solution B, reacting for 12h at 4 ℃, dialyzing and purifying for 3 days by using 0.02M PBS, and changing the solution three times every day to obtain the endosulfan-BSA conjugate which is the immunogen and is stored at-20 ℃ for later use.
3. A method for detecting the content of endosulfan in a sample comprises the following steps:
(1) Pretreating a sample;
(2) Detecting with the kit according to any one of claims 1 to 2;
(3) And analyzing the detection result.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010400219.6A CN111505297B (en) | 2020-05-13 | 2020-05-13 | Application of endosulfan artificial antigen in enzyme linked immunosorbent assay kit |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010400219.6A CN111505297B (en) | 2020-05-13 | 2020-05-13 | Application of endosulfan artificial antigen in enzyme linked immunosorbent assay kit |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN111505297A CN111505297A (en) | 2020-08-07 |
| CN111505297B true CN111505297B (en) | 2022-11-18 |
Family
ID=71869988
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202010400219.6A Active CN111505297B (en) | 2020-05-13 | 2020-05-13 | Application of endosulfan artificial antigen in enzyme linked immunosorbent assay kit |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN111505297B (en) |
Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1680406A (en) * | 2005-01-21 | 2005-10-12 | 浙江大学 | Triazoline artificial semi-antigen, antigen, specific antibody and use thereof |
| CN1700001A (en) * | 2005-02-24 | 2005-11-23 | 黄永 | Organic chlorine pesticide benzoepin artificial antigen and antibody, their preparation and use thereof |
| CN102939004A (en) * | 2010-04-06 | 2013-02-20 | 弗拉芒区生物技术研究所 | Specific delivery of agrochemicals |
| CN103601807A (en) * | 2013-04-18 | 2014-02-26 | 南京农业大学 | Preparation method of monoclonal antibody with oxime carbamate pesticide butocarboxim resistant property |
| CN106324243A (en) * | 2016-08-23 | 2017-01-11 | 广东志道医药科技有限公司 | Colloidal gold immunochromatography test strip and preparing and use method thereof |
| CN110133306A (en) * | 2019-05-09 | 2019-08-16 | 北京勤邦生物技术有限公司 | Detect enzyme linked immunological kit and its application of Cimaterol |
| CN110196322A (en) * | 2018-02-27 | 2019-09-03 | 广东志道医药科技有限公司 | Organophosphate and carbamate pesticide method for detecting residue and its test strips and preparation method |
Family Cites Families (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US10234470B2 (en) * | 2012-07-04 | 2019-03-19 | Beijing Kwinbon Biotechnology Co., Ltd. | Enzyme-linked immunosorbent assay kit for detecting dinitolmide and use thereof |
-
2020
- 2020-05-13 CN CN202010400219.6A patent/CN111505297B/en active Active
Patent Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1680406A (en) * | 2005-01-21 | 2005-10-12 | 浙江大学 | Triazoline artificial semi-antigen, antigen, specific antibody and use thereof |
| CN1700001A (en) * | 2005-02-24 | 2005-11-23 | 黄永 | Organic chlorine pesticide benzoepin artificial antigen and antibody, their preparation and use thereof |
| CN102939004A (en) * | 2010-04-06 | 2013-02-20 | 弗拉芒区生物技术研究所 | Specific delivery of agrochemicals |
| CN103601807A (en) * | 2013-04-18 | 2014-02-26 | 南京农业大学 | Preparation method of monoclonal antibody with oxime carbamate pesticide butocarboxim resistant property |
| CN106324243A (en) * | 2016-08-23 | 2017-01-11 | 广东志道医药科技有限公司 | Colloidal gold immunochromatography test strip and preparing and use method thereof |
| CN110196322A (en) * | 2018-02-27 | 2019-09-03 | 广东志道医药科技有限公司 | Organophosphate and carbamate pesticide method for detecting residue and its test strips and preparation method |
| CN110133306A (en) * | 2019-05-09 | 2019-08-16 | 北京勤邦生物技术有限公司 | Detect enzyme linked immunological kit and its application of Cimaterol |
Non-Patent Citations (1)
| Title |
|---|
| 环戊二烯类有机氯农药单克隆抗体的制备及鉴定;肖陈贵 等;《食品安全质量检测学报》;20141130;第5卷(第11期);第3425-3430页 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN111505297A (en) | 2020-08-07 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| AU2007257105B8 (en) | Elisa kit for detecting Sudan red and method thereof | |
| CN106324240B (en) | Detect enzyme linked immunological kit and its application of chlopyrifos | |
| CN110133306B (en) | Enzyme linked immunosorbent assay kit for detecting cimaterol and application thereof | |
| CN103288872A (en) | Methyl parathion hapten, and preparation method and application thereof | |
| CN104977406A (en) | Enzyme-linked immunoassay kit for detecting fluoroquinolone medicine and application of kit | |
| CN105675858B (en) | Detect enzyme linked immunological kit and its application of dichloro quinolinic acid | |
| CN105403703B (en) | Detect enzyme linked immunological kit and its application of carbendazim | |
| CN111812316B (en) | Application of fenpropathrin artificial antigen in enzyme linked immunosorbent assay kit | |
| CN103575885A (en) | Enzyme linked immunoassay kit for detecting T-2 toxin, and application thereof | |
| CN111505297B (en) | Application of endosulfan artificial antigen in enzyme linked immunosorbent assay kit | |
| CN109061171B (en) | ELISA kit for detecting flumetralin and application thereof | |
| CN101561439A (en) | Nitrofurantoin residue enzyme-linked immunoassay kit | |
| CN111848792B (en) | Preparation method and detection method of antibody of bovine serum albumin and other impurities in vaccine | |
| CN111505294B (en) | Application of artificial antigen of variolothricin in enzyme linked immunosorbent assay kit | |
| CN111896737B (en) | Application of snake-shaped bacterial artificial antigen in enzyme-linked immunosorbent assay kit | |
| CN113281501B (en) | Application of pentachloronitrobenzene artificial antigen in ELISA kit | |
| CN109265395B (en) | Preparation method and application of quinclorac hapten and antigen | |
| CN111443202B (en) | ELISA kit for detecting anticoagulant rodenticide, preparation and application thereof | |
| CN112904008A (en) | Enzyme linked immunosorbent assay kit for detecting protein A and other impurities in biological products and application thereof | |
| CN112098641B (en) | Application of inter-alternaria alternata artificial antigen in enzyme-linked immunosorbent assay kit | |
| CN105823872B (en) | Detect enzyme linked immunological kit and its application of maleic hydrazide | |
| CN109813922B (en) | Enzyme linked immunosorbent assay kit for detecting chlorpromazine and application thereof | |
| CN109735502B (en) | Hybridoma cell strain secreting salinomycin-resistant monoclonal antibody and application thereof | |
| CN112595850A (en) | Application of clothianidin artificial antigen in enzyme linked immunosorbent assay kit | |
| CN111812317A (en) | Application of polychlorinated biphenyl artificial antigen in enzyme linked immunosorbent assay kit |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |