CN106588699A - Isoprocarb hapten, artificial antigen and antibody and preparing method and application thereof - Google Patents
Isoprocarb hapten, artificial antigen and antibody and preparing method and application thereof Download PDFInfo
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
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- C07C271/00—Derivatives of carbamic acids, i.e. compounds containing any of the groups, the nitrogen atom not being part of nitro or nitroso groups
- C07C271/06—Esters of carbamic acids
- C07C271/40—Esters of carbamic acids having oxygen atoms of carbamate groups bound to carbon atoms of six-membered aromatic rings
- C07C271/42—Esters of carbamic acids having oxygen atoms of carbamate groups bound to carbon atoms of six-membered aromatic rings with the nitrogen atoms of the carbamate groups bound to hydrogen atoms or to acyclic carbon atoms
- C07C271/54—Esters of carbamic acids having oxygen atoms of carbamate groups bound to carbon atoms of six-membered aromatic rings with the nitrogen atoms of the carbamate groups bound to hydrogen atoms or to acyclic carbon atoms to carbon atoms of hydrocarbon radicals substituted by carboxyl groups
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- C07C201/00—Preparation of esters of nitric or nitrous acid or of compounds containing nitro or nitroso groups bound to a carbon skeleton
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- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/76—Albumins
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- C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
- C07K16/44—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material not provided for elsewhere, e.g. haptens, metals, DNA, RNA, amino acids
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- G01N33/531—Production of immunochemical test materials
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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
- G01N33/577—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
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- G01N2430/00—Assays, e.g. immunoassays or enzyme assays, involving synthetic organic compounds as analytes
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Abstract
The invention discloses an isoprocarb hapten, artificial antigen and antibody and a preparing method and application thereof. The structural formula of the isoprocarb hapten is shown in the formula (I), and the structural formula of the isoprocarb artificial antigen is shown in the formula (II). The linearity range of the antibody prepared based on the hapten and the artificial antigen is 1.88-82.8 ng/mL, the lowest detectable limit of the antibody is 0.54 ng/mL, and the hemi-inhibitory concentration of the antibody is 11.7 ng/mL. The specificity, sensitivity and accuracy of the antibody are high, and the antigen and antibody have a wide application prospect in the aspect of residue detection of isoprocarb in agricultural products. The formula is defined in the description.
Description
Technical field
The present invention relates to technical field of food safety detection, more particularly, to a kind of Mobucin hapten, artificial anti-
Former, antibody and its preparation method and application.
Background technology
Mobucin (Isoprocarb, MIPC) also known as go out and flutter scattered, isoprocarb, molecular formula is C11H15NO2, chemical name is 2-
Isopropyl phenyl-N- methyl carbamates, are the one kind in carbamate insecticidess, and its structural formula is as shown in (III).
Mobucin can pass through the biological activity for suppressing the acetylcholinesterase in insecticide body, so as to insect paralysis be caused
Extremely, all it is widely used in fruit and vegerable, grain, Nicotiana tabacum L. etc..But this kind of pesticide also has stronger bio-toxicity.
Mobucin detection method common at present mainly has high performance liquid chromatography (HPLC), gas chromatography (GC), height
Effect liquid phase chromatogram-MS (HPLC-MS) and GC-MS (GC-MS), but these detection methods are all
There is the defects such as instrument and equipment costliness, complex pretreatment, round of visits length, it is impossible to meet the quick detection of batch samples.Cause
This, it is necessary to develop simpler fast and easily detection method.
Immunologic detection method is one of important method of current food safety quick detection, with sensitivity height, specificity
By force, pre-treatment is simple, round of visits is short, the advantages of detect suitable for batch samples, research worker is developed using the method
The quick inspection product of multi-medicament, and the accreditation in market has been obtained, however, not yet occurring the immunity for Mobucin at present
The correlation technique report of detection method and product.
The content of the invention
The technical problem to be solved in the present invention is the deficiency for the existing detection technique of Mobucin, there is provided a kind of Mobucin half
Antigen.
The invention solves the problems that another technical problem be to provide the haptenic preparation method of the Mobucin.
A present invention also technical problem to be solved is to provide Mobucin artificial antigen.
The present invention is while the technical problem to be solved is to provide the application of the Mobucin artificial antigen.
A present invention also technical problem to be solved is to provide the preparation method of the Mobucin artificial antigen.
A present invention also technical problem to be solved is to provide the application of Mobucin antibody and antibody.
The purpose of the present invention is achieved by the following technical programs：
A kind of Mobucin hapten is provided, its structural formula is as shown in formula I：
Wherein, n is-CH2Group numbers, n=1～6.
Invention also provides the haptenic preparation method of the Mobucin, comprises the following steps：
S01. 2- isopropyl-phenols are dissolved in into dichloromethane, add pyridine to form mixed solution, under ice bath, will be to nitro
Benzene chloro-formate is added in the mixed solution, reaction；
S02. after the reaction described in step S01 terminates, washing reaction product takes organic faciess and is dried, and vacuum distillation is obtained
To product dissolved with solvent；Obtain solution 1；
4-Aminobutanoicacid is dissolved in saturated sodium bicarbonate solution, solution 2 is obtained；
S03. in ice-water bath, solution 1 is added in solution 2 and is reacted, after reaction terminates, adjust reaction gained reactant liquor
PH value is subacidity, is extracted with organic solvent, takes organic faciess drying, crosses column purification, collects target components, and vacuum distillation removes molten
Agent is obtained final product.
Preferably, it is using the side of Deca p-nitrophenyl chloroformate to be added in the mixed solution described in step S01
Formula.The ice bath preferably adopts 4 DEG C of ice salt baths.
Preferably, washing is to adopt dilute hydrochloric acid solution described in step S02；Further preferably described dilute hydrochloric acid solution it is dense
Spend for 1～6M.
Preferably, the drying adopts anhydrous sodium sulfate；
Preferably, solvent described in step S02 is tetrahydrofuran；
Preferably, the pH value for reactant liquor being adjusted described in step S03 adopts hydrochloric acid solution；
Preferably, organic solvent described in step S03 is ethyl acetate；The drying adopts anhydrous sodium sulfate.
Preferably, the concentration of hydrochloric acid solution described in step S03 is 4mol/L；The pH value is 6；
Preferably, the mol ratio of 2- isopropyl-phenols and Nitrophenyl chloroformate described in step S01 is 1:1.2～1:2, institute
The time for stating reaction is 2～5h.
Preferably, the mol ratio of product described in step S02 and 4-Aminobutanoicacid is 1:1.5～1:10.
Preferably, it is using 200～300 mesh silica column purifications that column purification is crossed described in step S03.
The present invention provides a kind of Mobucin artificial antigen, and its structural formula is as shown in formula II：
Wherein, n is-CH2Group numbers, n=1～6.
Present invention simultaneously provides application of the Mobucin artificial antigen in Mobucin antibody or detection Mobucin is prepared.
Present invention simultaneously provides the preparation method of the Mobucin artificial antigen, is to adopt active ester method by the Mobucin
Hapten (I) prepare with carrier protein couplet.
Specifically, the preparation method of the Mobucin artificial antigen, comprises the following steps：
S11. Mobucin hapten of the present invention is dissolved in DMF (DMF) to be formed and mixes molten
Liquid, adds 1,3- dicyclohexylcarbodiimides and N-hydroxy-succinamide in the mixed solution, stirring reaction under room temperature
Overnight, supernatant is taken after centrifugation；
S12. step S11 gained supernatant is added in carbonate (pH9.6) buffer solution of carrier protein, is stirred under room temperature
Reaction is mixed, after reaction terminates, the dialysis of gained reactant liquor the Mobucin artificial antigen is obtained into.
Preferably, Mobucin hapten described in step S11 and the mass ratio of carrier protein are 1:10～1:60.
Preferably, the carrier protein is that bovine serum albumin (BSA), ovalbumin (OVA), human serum albumin or blood are blue
Albumen.
Preferably, the time reacted described in step S12 is 8h.
Present invention simultaneously provides the Mobucin antibody prepared using the Mobucin artificial antigen, and the antibody
Application in detection Mobucin.
The invention has the advantages that：
The present invention provides first a kind of new Mobucin hapten, can be simple based on the Mobucin hapten
Prepare Mobucin artificial antigen and specific antibody, the range of linearity of the antibody is 1.88～82.8ng/mL, minimum inspection
Survey is limited to 0.54ng/mL, and 503nhibiting concentration is 11.7ng/mL, and the antibody specificity, sensitivity, accuracy are higher, can be very
Be applied to well to set up Mobucin and obtain immuno analytical method, overcome existing Mobucin residual instrument detection method apparatus expensive,
The defects such as complex pretreatment, round of visits length, so as to realize quick detection agricultural product in Mobucin residual, with wide application
Prospect.
Description of the drawings
Fig. 1 is the flow chart of the haptenic preparation technology of Mobucin.
Fig. 2 Mobucin hapten mass spectruies.
The flow chart of the preparation technology of Fig. 3 Mobucin artificial antigenes.
The ELISA competition test curve charts of Fig. 4 Mobucin antibody.
Specific embodiment
Further illustrate the inventive method with specific embodiment below in conjunction with the accompanying drawings.Following embodiments and accompanying drawing are only used for showing
Example property explanation, it is impossible to be interpreted as limitation of the present invention.Unless stated otherwise, the reagent raw material used in following embodiments is normal
The raw reagent raw material that commercial or commercial sources are obtained is advised, unless stated otherwise, the method and apparatus used in following embodiments is
Method and apparatus commonly used in the art.
The haptenic preparation method of the Mobucin of embodiment 1：
The haptenic preparation method course of reaction of Mobucin is as shown in Figure 1.
Comprise the following steps that：
S01. 1.36g 2- isopropyl-phenols are weighed to be dissolved in after 20mL dichloromethane, adds 0.5mL pyridines to form mixing molten
Liquid, under 4 DEG C of ice salt baths, in Deca 2.0g p-nitrophenyl chloroformate to mixed solution, reacts 3h；
S02. react step S01 gained reactant liquor with the dilute hydrochloric acid that molar concentration is 6M to wash, take organic faciess use
Anhydrous Na2SO4It is dried, then carries out vacuum distillation；Take product obtained above (0.67g) to be dissolved in 12mL tetrahydrofurans, obtain molten
Liquid 1；0.3g (2.4mmol) 4-Aminobutanoicacid is dissolved in 5mL saturated sodium bicarbonate solutions, solution 2 is obtained；
S03. in ice-water bath, solution 1 is added and react in solution 2 after 4.5h, Deca 4mol/L is continued in reactant liquor
HCl solution, the pH value of reactant liquor is adjusted to into subacidity, be extracted with ethyl acetate 3 times, take organic addition anhydrous Na2SO4It is dried,
With 200～300 mesh silica column purifications, Mobucin hapten is obtained, Mobucin hapten mass spectrum is as shown in Figure 2.
The preparation method of the Mobucin artificial antigen of embodiment 2
As shown in Figure 3, the preparation method of the present embodiment offer Mobucin immunogen/coating antigen is as follows：
The present embodiment immunogen is carrier protein with the preparation difference of coating antigen, and the immunogenic carrier albumen is adopted
With bovine serum albumin (BSA), the coating antigen carrier protein is using ovalbumin (OVA).Described below is with immunogenic preparation
As a example by method.
100 μm of gained Mobucin hapten of ol embodiments 1 are taken, in being dissolved in 0.5mL DMF, is then added in the solution
Equimolar positive tri-n-butylamine and ethyl chloroformate, allow it to be stirred at room temperature overnight after reaction and are centrifuged, take supernatant and be slowly added to
To in the carbonate buffer solution of the carbonate (pH9.6) of the carrier protein of 5mL 15mg/mL, then stirring reaction 8h, treats anti-
After the completion of answering, then reactant liquor is loaded into bag filter, dialysed 3 days with 0.01mol/L PBS (pH 7.4) buffer solution, changed daily
3 dialysis solution, are then centrifuged 5min by dialysis solution in 4000 turns/min, supernatant liquid are taken in -20 DEG C of preservations, for immunity.
The preparation and identification of the antibody of embodiment 3
The present invention is illustrated by taking the preparation method of monoclonal antibody as an example, according to inventive concept, people in the art
Member is referred to prior art and prepares polyclonal antibody and genetic engineering antibody.
The preparation method of monoclonal antibody is as follows：The gained immunogen of embodiment 2 is mixed into (with isodose immunological adjuvant
1 immunity Freund's complete adjuvant, later booster immunization is with incomplete Freund's adjuvant), after the complete emulsifying of agitator, abdominal part
The week old Balb/c female mice of subcutaneous injection 7～8, after the 4th booster immunization, utilizes later booster immunization every three weeks once
Indirect ELISA determines serum titer and suppression ratio, and choosing all higher mice of potency and suppression ratio carries out cell fusion, before fusion
3 days doubling dosage reinforced immunologicals are once.Cell fusion is carried out with mouse myeloma SP2/0 cells, hybridoma is obtained, will be miscellaneous
Hand over oncocyte to be placed in female Balb/c mices culturing in vivo, obtain the ascites containing high-concentration monoclonal antibody.Using octanoic acid-sulphuric acid
Ammonium salting out method carries out the monoclonal antibody that purification obtains high specific to ascites.
The application of the antigen of embodiment 4 and antibody and the foundation of enzyme-linked immunoassay method
The working concentration of antigen and antibody is determined by square formation titrimetry, the working concentration of coating antigen is 0.5 μ g/mL, different
The extension rate of the third prestige antibody is 1/40000.
Do experimental solutions with the Mobucin solution of variable concentrations, with 19.2ng/mL as its maximum concentration, gradient dilution under enter
Row competition, using 9 groups of parallel laboratory tests (n=3).
Indirect Competitive ELISA method：It is with the coated elisa plate of above-mentioned working concentration, experimental solutions are same with antibody-solutions
When be added in ELISA Plate aperture, while arranging blank well and negative control hole, 37 DEG C of incubation 40min pour out liquid in hole, use
Cleaning mixture is washed 5 times, ELISA Plate is upside down in absorbent paper and is patted：The ELIAS secondary antibody solution that addition has diluted, 37 DEG C of incubations
20min, is washed 5 times with cleaning mixture, is patted dry：Substrate chromophoric solution, 37 DEG C of incubation 10min is added to add terminate liquid color development stopping,
Light absorption value OD is determined at wavelength 450nm with microplate reader, with light absorption value OD as vertical coordinate, with the log10 of ICP experimental solutions concentration
It is worth for abscissa, draws semilog canonical plotting, sees Mobucin ELISA competition test curves shown in accompanying drawing 3.
As a result show that standard curve has complete reverse-s shape shape, and have upper mounting plate and lower platform, standard curve it is parallel
Measure number of times 3 times, experimental repeatability is good, and relative standard deviation is but within 15%.
Lowest detectable limit (LOD value) of the Mobucin antibody in buffer solution is calculated for 0.54ng/mL, half by formula
Amount of suppression (IC50Value) it is 11.7ng/mL, the range of linearity is 1.88～82.8ng/mL.Illustrate present invention may determine that obtaining hapten
With antigen and antibody, and in the detection of Mobucin, with preferably accuracy and sensitivity, can perform well in agricultural product and
The detection of Mobucin in food samples.
Embodiment described above only expresses the several embodiments of the present invention, and its description is more concrete and detailed, but and
Therefore the restriction to the scope of the claims of the present invention can not be interpreted as.It should be pointed out that for one of ordinary skill in the art
For, without departing from the inventive concept of the premise, some deformations and improvement can also be made, these belong to the guarantor of the present invention
Shield scope.Therefore, the protection domain of patent of the present invention should be defined by claims.
Claims (10)
1. a kind of Mobucin hapten, it is characterised in that its structural formula is as shown in formula I：
Wherein, n is-CH2Group numbers, n=1～6.
2. the haptenic preparation method of Mobucin described in a kind of claim 1, it is characterised in that comprise the following steps：
S01. 2- isopropyl-phenols are dissolved in into dichloromethane, add pyridine to form mixed solution, under ice bath, by p-nitrophenyl chlorine
Formic acid esters are added in the mixed solution, reaction；
S02. after the reaction described in step S01 terminates, washing reaction product takes organic faciess and is dried, and vacuum distillation is obtained
Product solvent dissolves；Obtain solution 1；
4-Aminobutanoicacid is dissolved in saturated sodium bicarbonate solution, solution 2 is obtained；
S03. in ice-water bath, solution 1 is added in solution 2 and is reacted, after reaction terminates, adjust the pH value of reaction gained reactant liquor
For subacidity, extracted with organic solvent, take organic faciess drying, cross column purification, collect target components, vacuum distillation removes solvent and is
；
Washing is to adopt hydrochloric acid solution described in preferred steps S02；The drying adopts anhydrous sodium sulfate；
Solvent described in preferred steps S02 is tetrahydrofuran；
The pH value that reactant liquor is adjusted described in preferred steps S03 adopts hydrochloric acid solution；
Organic solvent described in preferred steps S03 is ethyl acetate；The drying adopts anhydrous sodium sulfate.
3. the haptenic preparation method of Mobucin according to claim 2, it is characterised in that will be right described in preferred steps S01
It is to adopt Deca that Nitrophenyl chloroformate is added in the mixed solution；The pH of reaction gained reactant liquor is adjusted described in step S03
It is to adopt hydrochloric acid solution to be worth for subacidity；It is preferred that the concentration of the hydrochloric acid solution is 4mol/L；It is preferred that the pH value is 6.
4. the haptenic preparation method of Mobucin according to claim 2, it is characterised in that 2- isopropyls described in step S01
The mol ratio of phenol and Nitrophenyl chloroformate is 1:1.2～1:2；The time of the reaction is 2～5h；Produce described in step S02
The mol ratio of thing and 4-Aminobutanoicacid is 1:1.5～1:10.
5. a kind of Mobucin artificial antigen, it is characterised in that its structural formula is as shown in formula II：
Wherein, n is-CH2Group numbers, n=1～6.
6. application of the Mobucin artificial antigen described in claim 5 in Mobucin antibody or detection Mobucin is prepared.
7. the preparation method of Mobucin artificial antigen described in claim 5, it is characterised in that being will be described different using active ester method
Third prestige hapten (I) is prepared with carrier protein couplet.
8. the preparation method of Mobucin artificial antigen described in claim 7, it is characterised in that comprise the following steps：
S11. the Mobucin hapten described in any one of Claims 1-4 is dissolved in DMF and forms mixed
Solution is closed, 1,3- dicyclohexylcarbodiimides and N-hydroxy-succinamide are added in the mixed solution, stirred under room temperature
Reaction overnight, takes supernatant after centrifugation；
S12. step S11 gained supernatant is added in the carbonate buffer solution of carrier protein, stirring reaction under room temperature, reaction
After end, the dialysis of gained reactant liquor is obtained into the Mobucin artificial antigen.
9. the preparation method of Mobucin artificial antigen according to claim 8, it is characterised in that Mobucin described in step S11
Hapten is 1 with the mass ratio of carrier protein:10～1:60；It is Sanguis Bovis seu Bubali that carrier protein described in step S12 is the carrier protein
Albumin, ovalbumin, human serum albumin or hemocyanin；The time reacted described in step S12 is 8h.
10. Mobucin artificial antigen described in a kind of employing claim 5 prepares Mobucin monoclonal antibody, Anti-TNF-α
Body or genetic engineering antibody；Application of the antibody in detection Mobucin.
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Cited By (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107449900A (en) * | 2017-06-07 | 2017-12-08 | 广东产品质量监督检验研究院 | Carbamate and chrysanthemum esters multi-residue determination antigen, antibody and preparation method and application |
| CN107462715A (en) * | 2017-07-31 | 2017-12-12 | 深圳市药品检验研究院（深圳市医疗器械检测中心） | A kind of carbofuran and Mobucin duplex inspection Immunofluorescence test paper strip and kit and its application |
| CN109239336A (en) * | 2018-09-19 | 2019-01-18 | 北京勤邦生物技术有限公司 | A kind of test strips and its application detecting Mobucin |
| CN110423729A (en) * | 2019-06-27 | 2019-11-08 | 江南大学 | One plant of hybridoma cell strain GTY for secreting anti-Mobucin monoclonal antibody and its application |
| CN110734497A (en) * | 2019-10-25 | 2020-01-31 | 华南农业大学 | single-chain antibody for directly recognizing isoprocarb and preparation method and application thereof |
| CN112250617A (en) * | 2020-10-14 | 2021-01-22 | 华南农业大学 | Sodium picosulfate hapten, artificial antigen, antibody and preparation method and application thereof |
| CN113150162A (en) * | 2021-03-11 | 2021-07-23 | 广东康辉集团有限公司 | Preparation method and application of antibody of carbamate pesticide |
| CN113861078A (en) * | 2021-12-06 | 2021-12-31 | 信达安检测技术(天津)有限公司 | New isoprocarb hapten and complete antigen as well as preparation and application thereof |
Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPH08231591A (en) * | 1995-02-28 | 1996-09-10 | Otsuka Chem Co Ltd | Hapten, antigen, antibody specific to carbamate agent and detection of carbamate agent |
| JPH09176197A (en) * | 1995-12-26 | 1997-07-08 | Kankyo Meneki Gijutsu Kenkyusho:Kk | Antibody specific to 2-ethylthiomethylphenyl methyl carbamate and its production |
| CN1616481A (en) * | 2003-11-13 | 2005-05-18 | 南京农业大学 | Synthetic method for metolcarb artificial antigen |
| CN101407480A (en) * | 2008-11-21 | 2009-04-15 | 华南农业大学 | Preparation of semicarbazide derivative hapten, antigen and antibody |
| CN105137009A (en) * | 2015-09-18 | 2015-12-09 | 北京勤邦生物技术有限公司 | ELISA (enzyme linked immunosorbent assay) kit for detecting carbofuran and application of ELISA kit |
-
2016
- 2016-11-30 CN CN201611086555.8A patent/CN106588699A/en active Pending
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPH08231591A (en) * | 1995-02-28 | 1996-09-10 | Otsuka Chem Co Ltd | Hapten, antigen, antibody specific to carbamate agent and detection of carbamate agent |
| JPH09176197A (en) * | 1995-12-26 | 1997-07-08 | Kankyo Meneki Gijutsu Kenkyusho:Kk | Antibody specific to 2-ethylthiomethylphenyl methyl carbamate and its production |
| CN1616481A (en) * | 2003-11-13 | 2005-05-18 | 南京农业大学 | Synthetic method for metolcarb artificial antigen |
| CN101407480A (en) * | 2008-11-21 | 2009-04-15 | 华南农业大学 | Preparation of semicarbazide derivative hapten, antigen and antibody |
| CN105137009A (en) * | 2015-09-18 | 2015-12-09 | 北京勤邦生物技术有限公司 | ELISA (enzyme linked immunosorbent assay) kit for detecting carbofuran and application of ELISA kit |
Non-Patent Citations (6)
| Title |
|---|
| CAS登记号：1902704-92-0: "CAS登记号：1902704-92-0", 《STN REGISTRY数据库》 * |
| MARI´A J. MORENO等: "Production of Monoclonal Antibodies to the N-Methylcarbamate Pesticide Propoxur", 《J. AGRIC. FOOD CHEM.》 * |
| 施海燕等: "液相色谱法测定抗体与对硫磷、异丙威及其竞争原的吸附常数", 《分析化学研究简报》 * |
| 杨耀军等: "西维因人工抗原的合成新方法", 《过程工程学报》 * |
| 贲亚琍等: "农药人工抗原合成方法的设计与优选", 《农药》 * |
| 贲亚琍等: "叶蝉散人工抗原中间产物2-异丙基苯甲酰氯的合成", 《江西农业大学学报》 * |
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| CN113150162A (en) * | 2021-03-11 | 2021-07-23 | 广东康辉集团有限公司 | Preparation method and application of antibody of carbamate pesticide |
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