CN110054576A - A kind of isopropyl methoxalamine chirality haptens, artificial antigen and antibody and its preparation method and application - Google Patents
A kind of isopropyl methoxalamine chirality haptens, artificial antigen and antibody and its preparation method and application Download PDFInfo
- Publication number
- CN110054576A CN110054576A CN201910300552.7A CN201910300552A CN110054576A CN 110054576 A CN110054576 A CN 110054576A CN 201910300552 A CN201910300552 A CN 201910300552A CN 110054576 A CN110054576 A CN 110054576A
- Authority
- CN
- China
- Prior art keywords
- isopropyl methoxalamine
- haptens
- chirality
- isopropyl
- solution
- 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.)
- Granted
Links
- 125000001449 isopropyl group Chemical group [H]C([H])([H])C([H])(*)C([H])([H])[H] 0.000 title claims abstract description 111
- 239000000427 antigen Substances 0.000 title claims abstract description 37
- 102000036639 antigens Human genes 0.000 title claims abstract description 37
- 108091007433 antigens Proteins 0.000 title claims abstract description 37
- 238000002360 preparation method Methods 0.000 title claims abstract description 22
- 102000014914 Carrier Proteins Human genes 0.000 claims abstract description 17
- 108010078791 Carrier Proteins Proteins 0.000 claims abstract description 17
- 238000001514 detection method Methods 0.000 claims abstract description 15
- DKIDEFUBRARXTE-UHFFFAOYSA-N 3-mercaptopropanoic acid Chemical compound OC(=O)CCS DKIDEFUBRARXTE-UHFFFAOYSA-N 0.000 claims abstract description 11
- 241001465754 Metazoa Species 0.000 claims abstract description 4
- KWYUFKZDYYNOTN-UHFFFAOYSA-M Potassium hydroxide Chemical compound [OH-].[K+] KWYUFKZDYYNOTN-UHFFFAOYSA-M 0.000 claims description 18
- ZMXDDKWLCZADIW-UHFFFAOYSA-N N,N-Dimethylformamide Chemical compound CN(C)C=O ZMXDDKWLCZADIW-UHFFFAOYSA-N 0.000 claims description 17
- 239000000243 solution Substances 0.000 claims description 17
- 238000006243 chemical reaction Methods 0.000 claims description 15
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 8
- 238000002156 mixing Methods 0.000 claims description 6
- 238000003756 stirring Methods 0.000 claims description 6
- 150000001718 carbodiimides Chemical class 0.000 claims description 5
- 239000011259 mixed solution Substances 0.000 claims description 5
- 235000019441 ethanol Nutrition 0.000 claims description 4
- 239000008363 phosphate buffer Substances 0.000 claims description 4
- 239000012460 protein solution Substances 0.000 claims description 4
- 108091003079 Bovine Serum Albumin Proteins 0.000 claims description 3
- 108010058846 Ovalbumin Proteins 0.000 claims description 3
- 229940098773 bovine serum albumin Drugs 0.000 claims description 3
- 229940092253 ovalbumin Drugs 0.000 claims description 3
- XCSGPAVHZFQHGE-UHFFFAOYSA-N alachlor Chemical compound CCC1=CC=CC(CC)=C1N(COC)C(=O)CCl XCSGPAVHZFQHGE-UHFFFAOYSA-N 0.000 claims description 2
- 150000001412 amines Chemical class 0.000 claims description 2
- 238000000746 purification Methods 0.000 claims description 2
- 210000002966 serum Anatomy 0.000 claims description 2
- 244000025254 Cannabis sativa Species 0.000 claims 1
- 239000003814 drug Substances 0.000 abstract description 6
- 230000035945 sensitivity Effects 0.000 abstract description 6
- 235000013305 food Nutrition 0.000 abstract description 4
- 230000006340 racemization Effects 0.000 abstract description 4
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 description 21
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 14
- VLKZOEOYAKHREP-UHFFFAOYSA-N n-Hexane Chemical compound CCCCCC VLKZOEOYAKHREP-UHFFFAOYSA-N 0.000 description 12
- 239000007788 liquid Substances 0.000 description 9
- 238000000034 method Methods 0.000 description 7
- 239000000047 product Substances 0.000 description 7
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 6
- 229960000583 acetic acid Drugs 0.000 description 6
- 239000002671 adjuvant Substances 0.000 description 6
- 230000015572 biosynthetic process Effects 0.000 description 6
- 238000003786 synthesis reaction Methods 0.000 description 6
- 241000283973 Oryctolagus cuniculus Species 0.000 description 5
- 239000000575 pesticide Substances 0.000 description 5
- 238000004809 thin layer chromatography Methods 0.000 description 5
- 238000002965 ELISA Methods 0.000 description 4
- 230000001804 emulsifying effect Effects 0.000 description 4
- 239000012362 glacial acetic acid Substances 0.000 description 4
- 230000003053 immunization Effects 0.000 description 4
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 description 3
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 description 3
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 3
- 239000003153 chemical reaction reagent Substances 0.000 description 3
- 230000008878 coupling Effects 0.000 description 3
- 238000010168 coupling process Methods 0.000 description 3
- 238000005859 coupling reaction Methods 0.000 description 3
- 230000000694 effects Effects 0.000 description 3
- 238000004817 gas chromatography Methods 0.000 description 3
- 238000004128 high performance liquid chromatography Methods 0.000 description 3
- 229910052739 hydrogen Inorganic materials 0.000 description 3
- 239000000463 material Substances 0.000 description 3
- WWZKQHOCKIZLMA-UHFFFAOYSA-N octanoic acid Chemical compound CCCCCCCC(O)=O WWZKQHOCKIZLMA-UHFFFAOYSA-N 0.000 description 3
- 230000001376 precipitating effect Effects 0.000 description 3
- 239000000377 silicon dioxide Substances 0.000 description 3
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 3
- HBAQYPYDRFILMT-UHFFFAOYSA-N 8-[3-(1-cyclopropylpyrazol-4-yl)-1H-pyrazolo[4,3-d]pyrimidin-5-yl]-3-methyl-3,8-diazabicyclo[3.2.1]octan-2-one Chemical class C1(CC1)N1N=CC(=C1)C1=NNC2=C1N=C(N=C2)N1C2C(N(CC1CC2)C)=O HBAQYPYDRFILMT-UHFFFAOYSA-N 0.000 description 2
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 2
- ZHNUHDYFZUAESO-UHFFFAOYSA-N Formamide Chemical compound NC=O ZHNUHDYFZUAESO-UHFFFAOYSA-N 0.000 description 2
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 description 2
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 2
- HEDRZPFGACZZDS-MICDWDOJSA-N Trichloro(2H)methane Chemical compound [2H]C(Cl)(Cl)Cl HEDRZPFGACZZDS-MICDWDOJSA-N 0.000 description 2
- 210000004369 blood Anatomy 0.000 description 2
- 239000008280 blood Substances 0.000 description 2
- 239000000872 buffer Substances 0.000 description 2
- 239000007853 buffer solution Substances 0.000 description 2
- 229910052799 carbon Inorganic materials 0.000 description 2
- 239000011248 coating agent Substances 0.000 description 2
- 238000000576 coating method Methods 0.000 description 2
- 239000012895 dilution Substances 0.000 description 2
- 238000010790 dilution Methods 0.000 description 2
- 239000003480 eluent Substances 0.000 description 2
- 238000002474 experimental method Methods 0.000 description 2
- 239000004009 herbicide Substances 0.000 description 2
- 239000001257 hydrogen Substances 0.000 description 2
- 230000036039 immunity Effects 0.000 description 2
- 238000002649 immunization Methods 0.000 description 2
- 238000003018 immunoassay Methods 0.000 description 2
- 238000005259 measurement Methods 0.000 description 2
- 238000011587 new zealand white rabbit Methods 0.000 description 2
- 230000005311 nuclear magnetism Effects 0.000 description 2
- 239000012071 phase Substances 0.000 description 2
- 230000003068 static effect Effects 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- 230000001988 toxicity Effects 0.000 description 2
- 231100000419 toxicity Toxicity 0.000 description 2
- 238000005160 1H NMR spectroscopy Methods 0.000 description 1
- WVQBLGZPHOPPFO-UHFFFAOYSA-N 2-chloro-N-(2-ethyl-6-methylphenyl)-N-(1-methoxypropan-2-yl)acetamide Chemical compound CCC1=CC=CC(C)=C1N(C(C)COC)C(=O)CCl WVQBLGZPHOPPFO-UHFFFAOYSA-N 0.000 description 1
- QCVGEOXPDFCNHA-UHFFFAOYSA-N 5,5-dimethyl-2,4-dioxo-1,3-oxazolidine-3-carboxamide Chemical compound CC1(C)OC(=O)N(C(N)=O)C1=O QCVGEOXPDFCNHA-UHFFFAOYSA-N 0.000 description 1
- VHUUQVKOLVNVRT-UHFFFAOYSA-N Ammonium hydroxide Chemical compound [NH4+].[OH-] VHUUQVKOLVNVRT-UHFFFAOYSA-N 0.000 description 1
- 239000005635 Caprylic acid (CAS 124-07-2) Substances 0.000 description 1
- 241001494246 Daphnia magna Species 0.000 description 1
- 108010000912 Egg Proteins Proteins 0.000 description 1
- 102000002322 Egg Proteins Human genes 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 239000007836 KH2PO4 Substances 0.000 description 1
- 239000007832 Na2SO4 Substances 0.000 description 1
- 241001597008 Nomeidae Species 0.000 description 1
- 238000010521 absorption reaction Methods 0.000 description 1
- 239000008351 acetate buffer Substances 0.000 description 1
- BIGPRXCJEDHCLP-UHFFFAOYSA-N ammonium bisulfate Chemical compound [NH4+].OS([O-])(=O)=O BIGPRXCJEDHCLP-UHFFFAOYSA-N 0.000 description 1
- 239000000908 ammonium hydroxide Substances 0.000 description 1
- BFNBIHQBYMNNAN-UHFFFAOYSA-N ammonium sulfate Chemical compound N.N.OS(O)(=O)=O BFNBIHQBYMNNAN-UHFFFAOYSA-N 0.000 description 1
- 229910052921 ammonium sulfate Inorganic materials 0.000 description 1
- 235000011130 ammonium sulphate Nutrition 0.000 description 1
- 238000011091 antibody purification Methods 0.000 description 1
- GEWYFWXMYWWLHW-UHFFFAOYSA-N azanium;octanoate Chemical compound [NH4+].CCCCCCCC([O-])=O GEWYFWXMYWWLHW-UHFFFAOYSA-N 0.000 description 1
- 230000000711 cancerogenic effect Effects 0.000 description 1
- 125000003178 carboxy group Chemical group [H]OC(*)=O 0.000 description 1
- 231100000357 carcinogen Toxicity 0.000 description 1
- 239000003183 carcinogenic agent Substances 0.000 description 1
- 239000007795 chemical reaction product Substances 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 238000004587 chromatography analysis Methods 0.000 description 1
- 238000001142 circular dichroism spectrum Methods 0.000 description 1
- 238000004140 cleaning Methods 0.000 description 1
- 238000004440 column chromatography Methods 0.000 description 1
- 230000002860 competitive effect Effects 0.000 description 1
- 239000002131 composite material Substances 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 230000007812 deficiency Effects 0.000 description 1
- 239000008367 deionised water Substances 0.000 description 1
- 229910021641 deionized water Inorganic materials 0.000 description 1
- UQLDLKMNUJERMK-UHFFFAOYSA-L di(octadecanoyloxy)lead Chemical compound [Pb+2].CCCCCCCCCCCCCCCCCC([O-])=O.CCCCCCCCCCCCCCCCCC([O-])=O UQLDLKMNUJERMK-UHFFFAOYSA-L 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 238000000502 dialysis Methods 0.000 description 1
- 239000003085 diluting agent Substances 0.000 description 1
- BNIILDVGGAEEIG-UHFFFAOYSA-L disodium hydrogen phosphate Chemical compound [Na+].[Na+].OP([O-])([O-])=O BNIILDVGGAEEIG-UHFFFAOYSA-L 0.000 description 1
- 229910000397 disodium phosphate Inorganic materials 0.000 description 1
- 238000006073 displacement reaction Methods 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- 230000000857 drug effect Effects 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 235000014103 egg white Nutrition 0.000 description 1
- 210000000969 egg white Anatomy 0.000 description 1
- 238000002330 electrospray ionisation mass spectrometry Methods 0.000 description 1
- 238000004945 emulsification Methods 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- 239000012467 final product Substances 0.000 description 1
- 238000005194 fractionation Methods 0.000 description 1
- 230000002363 herbicidal effect Effects 0.000 description 1
- 238000011534 incubation Methods 0.000 description 1
- 238000007689 inspection Methods 0.000 description 1
- 230000031700 light absorption Effects 0.000 description 1
- 238000004811 liquid chromatography Methods 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 238000013507 mapping Methods 0.000 description 1
- 238000001819 mass spectrum Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 238000012544 monitoring process Methods 0.000 description 1
- 229910000402 monopotassium phosphate Inorganic materials 0.000 description 1
- 238000006386 neutralization reaction Methods 0.000 description 1
- 229960002446 octanoic acid Drugs 0.000 description 1
- 230000003287 optical effect Effects 0.000 description 1
- 239000012074 organic phase Substances 0.000 description 1
- 230000000361 pesticidal effect Effects 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 239000002244 precipitate Substances 0.000 description 1
- 238000002203 pretreatment Methods 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 239000012266 salt solution Substances 0.000 description 1
- 238000005070 sampling Methods 0.000 description 1
- 229920006395 saturated elastomer Polymers 0.000 description 1
- 239000012047 saturated solution Substances 0.000 description 1
- 238000007789 sealing Methods 0.000 description 1
- 235000020183 skimmed milk Nutrition 0.000 description 1
- 239000011780 sodium chloride Substances 0.000 description 1
- 229910052938 sodium sulfate Inorganic materials 0.000 description 1
- 239000002904 solvent Substances 0.000 description 1
- 230000003595 spectral effect Effects 0.000 description 1
- 238000001228 spectrum Methods 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 238000001308 synthesis method Methods 0.000 description 1
- 238000010998 test method Methods 0.000 description 1
- 231100000027 toxicology Toxicity 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
- C07C321/00—Thiols, sulfides, hydropolysulfides or polysulfides
- C07C321/12—Sulfides, hydropolysulfides, or polysulfides having thio groups bound to acyclic carbon atoms
- C07C321/14—Sulfides, hydropolysulfides, or polysulfides having thio groups bound to acyclic carbon atoms of an acyclic saturated carbon skeleton
-
- 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
-
- 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
- C07K14/77—Ovalbumin
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
- C07K16/06—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies from serum
- C07K16/065—Purification, fragmentation
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- 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
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K19/00—Hybrid peptides, i.e. peptides covalently bound to nucleic acids, or non-covalently bound protein-protein complexes
-
- 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
- G01N33/531—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
-
- 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)
Landscapes
- Chemical & Material Sciences (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Organic Chemistry (AREA)
- Molecular Biology (AREA)
- Immunology (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- Medicinal Chemistry (AREA)
- Engineering & Computer Science (AREA)
- Biophysics (AREA)
- Genetics & Genomics (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Urology & Nephrology (AREA)
- Biomedical Technology (AREA)
- Hematology (AREA)
- Microbiology (AREA)
- Biotechnology (AREA)
- Cell Biology (AREA)
- Zoology (AREA)
- Gastroenterology & Hepatology (AREA)
- Toxicology (AREA)
- Food Science & Technology (AREA)
- Physics & Mathematics (AREA)
- Analytical Chemistry (AREA)
- General Physics & Mathematics (AREA)
- Pathology (AREA)
- Peptides Or Proteins (AREA)
Abstract
The invention discloses a kind of isopropyl methoxalamine chirality haptens, artificial antigen and antibody and its preparation method and application.The isopropyl methoxalamine chirality haptens has the molecular structure as shown in formula (I):The present invention first reacts racemization isopropyl methoxalamine with 3- mercaptopropionic acid, obtains haptens；This haptens is subjected to chiral resolution again, four stereoisomer haptens α RS-, α RR-, α SS-, α SR-MMPA is split as, artificial antigen then is made with carrier protein couplet respectively.This 4 kinds of artificial antigens are immunized to animal respectively and obtain enantioselectivity antibody, the IC that 4 kinds of obtained antibody detects its enantiomer of drugs50Respectively 28.89ng/mL, 13.34ng/mL, 27.07ng/mL, 5.68ng/mL, high specificity, high sensitivity can be used for the field quick detection of food safety, have broad application prospects.
Description
Technical field
The present invention relates to technical field of food safety detection, and in particular, to a kind of isopropyl methoxalamine chirality haptens, people
Work antigen and antibody and its preparation method and application.
Background technique
Isopropyl methoxalamine (Metolachlor, MET) is acetamide-group herbicides, is that China produces at present and usage amount is maximum
One of herbicide.There are two its tools chiral centre, chiral N axis and chiral carbon, thus, there are four types of stereoisomers for tool.It is removed
Careless activity is mainly contributed by middle S- type enantiomer, therefore there are two types of presently commercially available isopropyl methoxalamine Pesticidal products, one kind is to disappear outside
Product (Rac- type, also known as all thats) is revolved, another kind is S- anomeric product (S- type also known as Jin Douer).According to Environmental Protection Agency (US
EPA it) divides, isopropyl methoxalamine is classified as C class pesticide, is potential carcinogen matter.
Existing research shows that there are great differences in terms of toxicology with Rac- type isopropyl methoxalamine enantiomer for S- type, example
Such as, racemic isopropyl methoxalamine is 50~100 times of S- type isopropyl methoxalamine to the physiological-toxicity of Daphnia magna.It is main now to sell pesticide
It is to promote drug effect by improving the content of certain effective enantiomer, reduce toxicity, but due to the limitation of production cost and technology,
Raceme is still used now.Chiral pesticide bioactivity and toxicity have enantioselectivity, also do not there is corresponding limit standard, therefore
It is necessary to individually be tested and analyzed from enantiomer level.
At present the detection method of racemization isopropyl methoxalamine mainly have thin-layer chromatography (Thin Layer chromatography,
TLC), high performance liquid chromatography (High performance liquid chromatography, HPLC), gas-chromatography (Gas
Chromatography, GC), the methods of mass spectrum (Mass sepectrum, MS).Thin-layered chromatography remolding sensitivity is lower；Efficiently
Liquid chromatography and gas chromatography high sensitivity, measurement are accurate, are the main methods of current measurement isopropyl methoxalamine, but instrument
Equipment is expensive, and sample pre-treatments are complicated, and testing cost is high, high to experimenter's professional skill requirement.Inspection for chiral material
Survey, only by the above method cannot complete resolved detection go out, need by chiral column, chiral derivatizing reagents.And immunoassay
Method has the features such as quick, sensitive, accurate, easy to operate, and of less demanding to sample purity, especially suitable for batch samples
Detection.Therefore, to the detection of the chiral drug stereoisomer of realization, then need to prepare these isomeries of specific recognition
The antibody of body.
What is reported at present is mostly anti-racemization isopropyl methoxalamine antibody, has not been reported the preparation method of its chiral antibody, also not
It is detected in enantiomer level.
Summary of the invention
The technical problem to be solved by the present invention is to fail through immunoassay in enantiomer level for the prior art
Accurate detection isopropyl methoxalamine provides a kind of isopropyl methoxalamine chirality haptens to overcome the above-mentioned deficiency of the prior art, with
The artificial antigen of the haptens and carrier protein couplet preparation is immunogene, the specific isopropyl methoxalamine Anti-TNF-α of preparation
Body can quickly and effectively distinguish different isopropyl methoxalamine enantiomers, can be used for the field quick detection of food safety.
The object of the present invention is to provide a kind of isopropyl methoxalamine chirality haptens.
Another object of the present invention is to provide the preparation methods of above-mentioned isopropyl methoxalamine chirality haptens.
Another object of the present invention is to provide above-mentioned isopropyl methoxalamine chirality haptens to prepare isopropyl methoxalamine artificial
Application in antigen and/or antibody.
Another object of the present invention is to provide a kind of isopropyl methoxalamine artificial antigens.
Another object of the present invention is to provide the preparation methods of above-mentioned isopropyl methoxalamine artificial antigen.
Another object of the present invention is to provide a kind of isopropyl methoxalamine polyclonal antibodies.
Another object of the present invention is to provide above-mentioned isopropyl methoxalamine polyclonal antibodies to detect isopropyl methoxalamine in preparation
Application in the kit of enantiomer.
To achieve the goals above, the present invention is achieved by following scheme:
The present invention splits isopropyl methoxalamine derivative by chiral column, obtains 4 kinds of stereoisomer derivatives, i.e.,
For four kinds of isopropyl methoxalamine chirality haptens.Using the artificial antigen of the haptens and carrier protein couplet preparation as immunogene, system
Standby specific isopropyl methoxalamine polyclonal antibody, can quickly and effectively distinguish different isopropyl methoxalamine enantiomers, to respective
Chiral pesticide has preferable detection effect.
Therefore, a kind of isopropyl methoxalamine chirality haptens is claimed in the present invention, has the molecule knot as shown in formula (I)
Structure:
The preparation method of above-mentioned isopropyl methoxalamine chirality haptens is to use 3- sulfydryl third using isopropyl methoxalamine as object
Sour (3-MPA) is that derivating agent is specifically comprised the following steps: with one-step synthesis method isopropyl methoxalamine haptens (MMPA)
S1. 3- mercaptopropionic acid, isopropyl methoxalamine are dissolved in ethyl alcohol, add KOH, flowed back and stir under the conditions of 65~85 DEG C
Reaction 5h~8h is mixed, isopropyl methoxalamine derivative is obtained；
S2. isopropyl methoxalamine derivative is carried out by chiral resolution using AD-H column, obtains four kinds of stereoisomer derivatives, i.e.,
Obtain four kinds of isopropyl methoxalamine chirality haptens.
Preferably, the temperature of reaction described in step S1 is 80 DEG C, the time of reaction is 6h.
Preferably, isopropyl methoxalamine described in step S1,3- mercaptopropionic acid, potassium hydroxide molal weight ratio be 1:1:1
~4.
It is highly preferred that the molal weight ratio of isopropyl methoxalamine described in step S1,3- mercaptopropionic acid, potassium hydroxide is 1:1:
3。
The present invention be also claimed above-mentioned isopropyl methoxalamine chirality haptens prepare isopropyl methoxalamine artificial antigen and/or
Application in antibody.
A kind of isopropyl methoxalamine artificial antigen (MMPA-BSA) is by above-mentioned isopropyl methoxalamine chirality haptens and carrier egg
White coupling obtains.
Preferably, the carrier protein is bovine serum albumin(BSA) (BSA) or ovalbumin (OVA).
The preparation method of above-mentioned isopropyl methoxalamine artificial antigen is also claimed in the present invention, includes the following steps:
S1. above-mentioned isopropyl methoxalamine chirality haptens is dissolved in dimethyl formamide solution at room temperature, is mixed
After obtain isopropyl methoxalamine chirality haptens solution；
S2. carrier protein is dissolved in PBS solution and is uniformly mixed, obtain carrier protein solution；Again by isopropyl obtained by step S1
Alachlor chirality haptens solution is added dropwise in carrier protein solution, is obtained mixed solution, is stirred under the conditions of -4 DEG C, 100r/min
It mixes；
S3. carbodiimide is dissolved in dimethyl formamide solution, obtained Carbodiimide solution slowly drips while stirring
It adds in mixed solution obtained by step S2, in -4 DEG C of 8~12h of reaction；It is dialysed with phosphate buffer after reaction to get arriving
Isopropyl methoxalamine artificial antigen ɑ RS-MMPA-BSA, α RR-MMPA-BSA, α SS-MMPA-BSA, α SR-MMPA-BSA.
Preferably, the time of reaction described in step S3 is 9h.
Preferably, the molal weight ratio of the carrier protein and isopropyl methoxalamine chirality haptens is 1:50~100, diformazan
The volume fraction of base formamide is 10~15%.
It is highly preferred that the molal weight ratio of the carrier protein and isopropyl methoxalamine chirality haptens is 1:65.
Above-mentioned isopropyl methoxalamine artificial antigen is also claimed in preparing isopropyl methoxalamine polyclonal antibody in the present invention
Using.
A kind of isopropyl methoxalamine polyclonal antibody is animal to be immunized by above-mentioned isopropyl methoxalamine artificial antigen, then obtain dynamic
Object serum obtains after purification.
The preparation method of above-mentioned isopropyl methoxalamine polyclonal antibody, specifically includes the following steps:
With the artificial antigen obtained after 4 optical stereo isomers coupling carrier albumen of isopropyl methoxalamine carboxy derivatives
Immune new zealand white rabbit, respectively by the artificial antigen solution of 500 μ g/mL and equivalent adjuvant mixing and emulsifying, initial immunity and not
It being immunized after family name's Freund's complete adjuvant mixing and emulsifying, booster immunization later is immunized after being emulsified using immunogene and incomplete Freund's adjuvant,
It is immunized five times altogether, every minor tick 2 weeks, each immunizing dose is 1.2mL/；It takes a blood sample within 5th time immune latter tenth day, use is pungent
Acid-ammonium sulfate method purifies up to isopropyl methoxalamine polyclonal antibody.
The IC that gained isopropyl methoxalamine polyclonal antibody detects its enantiomer of drugs50Respectively 28.89ng/mL,
13.34ng/mL, 27.07ng/mL, 5.68ng/mL, high specificity, high sensitivity can quickly and effectively distinguish different isopropyl first
Careless amine enantiomer has preferable detection effect to respective Chiral pesticide.
Therefore, the present invention is also claimed above-mentioned isopropyl methoxalamine polyclonal antibody and detects isopropyl methoxalamine mapping in preparation
Application in the kit of body.
A kind of kit for detecting isopropyl methoxalamine enantiomer is also claimed in the present invention, and the kit includes above-mentioned different
Methoxalamine chirality haptens, isopropyl methoxalamine artificial antigen and/or isopropyl methoxalamine polyclonal antibody.
Compared with prior art, the invention has the following advantages:
(1) present invention first reacts racemization isopropyl methoxalamine with 3- mercaptopropionic acid, obtains haptens；Again by this haptens
Carry out chiral resolution, be split as four stereoisomer haptens α RS-, α RR-, α SS-, α SR-MMPA, then respectively with carrier
Artificial antigen is made in albumen coupling.This 4 kinds of artificial antigen difference immune rabbits are obtained into enantioselectivity antibody, 4 kinds obtained
The IC that antibody detects its enantiomer of drugs50Respectively 28.89ng/mL, 13.34ng/mL, 27.07ng/mL, 5.68ng/
ML, high specificity, high sensitivity can be used for the field quick detection of food safety, have broad application prospects.
(2) present invention reacts small molecule isopropyl methoxalamine with 3- mercaptopropionic acid, only needs single step reaction method that mesh can be obtained
Product is marked, yield is high, and it is easy to operate, it is suitable for large-scale promotion and application.
Detailed description of the invention
Fig. 1 is that isopropyl methoxalamine derivative synthesizes schematic diagram.
Fig. 2 is isopropyl methoxalamine derivative split result.
Fig. 3 is that isopropyl methoxalamine derivative identifies map.
Fig. 4 is that four kinds of artificial antigen UV scannings of isopropyl methoxalamine identify curve.
Fig. 5 is isopropyl methoxalamine indirect competitive ELISA standard curve.
Specific embodiment
With reference to the accompanying drawings of the specification and specific embodiment is made the present invention and is further elaborated, the embodiment
It is served only for explaining the present invention, be not intended to limit the scope of the present invention.Test method as used in the following examples is such as without spy
Different explanation, is conventional method；Used material, reagent etc., unless otherwise specified, for the reagent commercially obtained
And material.
The synthesis of 1 isopropyl methoxalamine haptens of embodiment
1, the synthesis of isopropyl methoxalamine haptens
Small haptens synthetic route chart is as shown in Figure 1, specific reaction process is as follows:
(1) the 3- mercaptopropionic acid (3-MPA) of 0.4g, 1g (about 1mL) isopropyl methoxalamine are added to and fill 50mL ethyl alcohol
In round-bottomed flask, the KOH of 0.66g is then added；Return stirring reacts 6h in thermostatical oil bath (80 DEG C)；
(2) thin layer chromatography is used, monitors reaction process every 2h；Solvent are as follows: methylene chloride adds two drop ammonium hydroxide, or
Be by n-hexane: ethyl acetate: acetic acid is the mixed solution of 2:1:0.1 by volume；
(3) it after fully reacting, is spin-dried for about 5mL, sucks in separatory funnel, a small amount of ethyl acetate is added and washes reaction flask；With
Saturated salt solution is extracted twice, and organic phase is poured into triangular flask A after water phase is 2~3 with HCl neutralization to pH and is extracted with ethyl acetate
It takes twice；Use Na2SO4Liquid is spin-dried for pico- wet by dry 1h；
(4) column chromatographs: pillar first being carried out sealing propertytest, cleaning, drying are spare after no leakage；To slightly wet rotation
It steams and 2~3 spoons of thick silica whites (80~100 mesh) is added in bottle, revolving is dry, becomes dried powder；Thin silica white is first added into column
(200~300 mesh), height are about pillar half, add the thick silica white of adsorbed product；Eluent is slowly added to pillar
In, eluent sequence are as follows: a. n-hexane: ethyl acetate: glacial acetic acid=80:10:1；B. n-hexane: ethyl acetate: glacial acetic acid=
40:10:0.5；C. n-hexane: ethyl acetate: glacial acetic acid=20:10:0.3；D. n-hexane: ethyl acetate: glacial acetic acid=10:
10:0.1；Column chromatography procedure carries out the monitoring reaction of TLC contact plate, liquid in the pipe for having product is combined into revolving bottle, in 37
DEG C revolving, 2h is spin-dried for, and obtains product.
2, it identifies
Isopropyl methoxalamine haptens is subjected to hydrogen spectrum nuclear-magnetism and Mass Spectrometric Identification.
It is as follows that hydrogen composes nuclear-magnetism result:1H-NMR (400MHz, CDCl3)
δH7.18-7.25 (2H, m, H-4 ', 5 '), 7.11 (1H, d, J=7.2Hz, H-3 '), 4.16-4.20 (1H, m, H-
5), 3.67-3.76 (1H, m, H-6a), 3.45-3.49 (1H, m, H-6b), 3.27 (3H, d, J=9.6Hz, H-7), 2.87-
2.90 (4H, m, H-3,4), 2.65 (2H, t, J=7.2Hz, H-2), 2.53-2.62 (2H, m, C2’-CH2-),2.25(3H,d,J
=9.2Hz, C5-CH3), 1.25 (3H, t, J=7.2Hz, C6’-CH3), 1.15 (3H, t, J=6.8Hz, C2’-CH3)
Mass spectral results are as follows: ESI-MS:m/z [M-H]-352.2(C18H27NO4S)。
The two result all shows composite result correctly and successfully, and successfully synthesis obtains isopropyl methoxalamine hapten derivant knot
Structure.
2 haptens of embodiment is split and identification
1, the fractionation of haptens
Isopropyl methoxalamine derivative is split using CHIRALPAK AD-H chiral column, mobile phase n-hexane: ethyl alcohol:
Acetic acid=90:10:0.1, flow velocity 1mL/min, are detected at 220nm by 35 DEG C of column temperature, and it is as shown in Figure 2 to split spectrogram.
2, the identification of enantiomer
Four kinds of haptens are dissolved with acetonitrile, are each configured to 1mg/mL solution.It is surveyed using Chirascan circular dichroism spectra
It is fixed, it is scanned at 190~300nm, bandwidth 1nm, stepping 1nm, sampling time 0.5s, obtains experiment ECD spectrogram.Pass through Gauss again
Software calculates, and obtains calculating map, experiment and the comparison of calculating map are as shown in figure 3, peak sequence can be determined are as follows: α RS, α
RR, α SS, α SR.
The synthesis and identification of 3 artificial antigen of embodiment
1, the synthesis of artificial antigen
(1) 3.5mg isopropyl methoxalamine is accurately weighed, 200 μ L dimethylformamides (DMF) are dissolved in, is A liquid；
(2) 10mg bovine serum albumin(BSA) (BSA) is added in the PBS (0.1moL/L, pH 7.4) of 1.7mL, is B liquid；
(3) 4.22mg carbodiimide is dissolved in 100 μ L dimethylformamides (DMF), is C liquid；
(4) B liquid is added in A drop, C liquid is then slowly added dropwise, become edged stirring, -4 DEG C of reaction 9h；
(5) it is dialysed two days with phosphate buffer, 4 times a day, racemization isopropyl methoxalamine artificial antigen is obtained after dialysis.
Phosphate buffer (0.01M, pH7.4): Na2HPO4·12H2O 2.90g, NaCl 8.50g, KCl 0.20g,
KH2PO40.20g adds deionized water to be settled to 1000mL.
2, it identifies
The artificial antigen of above-mentioned synthesis is taken to carry out UV scanning, as a result as shown in Figure 4.
Isopropyl methoxalamine, BSA and artificial antigen are carried out to ultraviolet (200~400nm) scanning identification respectively, and by comparing
It is coupled the highest light absorption value of each substance of front and back, it is found that the absorption curve of artificial antigen is significantly different with carrier protein, BSA only exists
There is characteristic peak at 280nm, comparison three's curve, which can be seen that, to be subjected to displacement, therefore illustrates that reaction product is answering for carrier protein and MMPA
Object is closed, is coupled successfully.
The preparation and purifying of 4 isopropyl methoxalamine enantioselectivity polyclonal antibody of embodiment
1, four kinds of stereoisomer polyclonal antibody preparations
New zealand white rabbit is immunized with four kinds of artificial antigens of isopropyl methoxalamine, by the artificial antigen solution of 500 μ g/mL with
It is immunized after equivalent adjuvant mixing and emulsifying, initial immunity and Freund's complete adjuvant mixing and emulsifying, booster immunization later uses immune
It being immunized, is immunized five times altogether, every minor tick 2 weeks after the emulsification of former and incomplete Freund's adjuvant, each immunizing dose is 1.2mL/, the
It takes a blood sample within five times immune latter tenth day.
2, isopropyl methoxalamine polyclonal antibody purification (caprylic acid-ammonium)
(1) rabbit anteserum is taken, under the conditions of 4 DEG C, 10000r/min is centrifuged 10min, removal precipitating；
(2) take the rabbit anteserum of 1 volume to mix (pH4.5) with the acetate buffer of 2 volumes, be stirred at room temperature down, dropwise plus
Enter caprylic acid, usage amount is 75 μ L caprylic acids/mL rabbit anteserum；
(3) mixing 30min, 4 DEG C of static 2h, which is stirred at room temperature, precipitates it sufficiently；
Under the conditions of (4) 4 DEG C, 10000r/min is centrifuged 10min, removal precipitating；
(5) PBS buffer solution (0.1M, pH7.4) of 1/10 volume is added, with the sodium hydroxide tune pH to 7.4 of 2M, calculates total
Liquor capacity；
(6) 45% saturated solution is become in the ammonium sulfate that 0.277g/mL is added in 30min under ice bath；
After (7) 4 DEG C of static 1h or more, under the conditions of 4 DEG C, 10000r/min is centrifuged 10min, abandons supernatant；
(8) after precipitating is dissolved and is dialysed three days with PBS buffer solution (0.1M, pH 7.4) to obtain the final product.
The ELISA of the anti-isopropyl methoxalamine antibody of embodiment 5 is detected
1, ELISA detection method
(1) Rac-MMPA-OVA artificial antigen is diluted to the concentration of 0.25 μ g/mL with coating buffer, is coated with 96 hole enzyme marks
100 μ L are added in plate, every hole, and 37 DEG C are incubated overnight；
(2) coating buffer is discarded, is washed 2 times；
(3) 120 μ L confining liquids (5% skimmed milk power), 37 DEG C of closing 3h are added in every hole；
(4) confining liquid is discarded, is had the final say, 37 DEG C are dried for standby；
(5) isopropyl methoxalamine polyclonal antibody is diluted with PBST 1:16000 times, and isopropyl methoxalamine MET is diluted to
10000,1000,100,10,1,0.1,0.01ng/mL；
(6) every row adds 50 μ L isopropyl methoxalamine dilutions (three groups are parallel), then plus 50 μ L to antibody diluent hole, at 37 DEG C
40min is incubated, is washed 5 times；
(7) the anti-HRP of goat-anti rabbit two (5000 times of dilutions) is added, 37 DEG C of incubation 30min are washed 5 times, clappers；
(8) developing solution colour developing 10min is added；
(9) 50 μ L 10%H are added2SO4Reaction is terminated, and reads OD value at 450nm.
2, testing result
The immune gained antibody indirect of α RS-MMPA-BSA, α RR-MMPA-BSA, α SS-MMPA-BSA, α SR-MMPA-BSA is competing
It is as shown in Figure 5 to strive ELISA standard curve.
Its IC50Respectively 28.89ng/mL, 13.34ng/mL, 27.07ng/mL, 5.68ng/mL.
Detection limit (LOD) is 0.86ng/mL, 0.85ng/mL, 0.91ng/mL, 0.22ng/mL.
IC20-IC80Respectively 4.18~199.67ng/mL, 1.57~113.61ng/mL, 5.18~141.41ng/mL,
0.44~73.02ng/mL.
For anti alpha RS-MET antibody to α RR-MET, the cross reacting rate of α SS-MET, α SR-MET are respectively 2.94%,
27.96%, 4.75%.
Anti alpha RR-MET antibody is 0.42%, 0.68% to α RS-MET, the cross reacting rate of α SS-MET, α SR-MET,
1.67%.
Anti alpha SS-MET antibody is 57.25%, 5.27% to α RS-MET, the cross reacting rate of α RR-MET, α SR-MET,
1.2%.
Anti alpha SR-MET antibody is respectively as follows: 8.67% to α RS-MET, the cross reacting rate of α RR-MET, α SS-MET,
23.48%, 4.35%.
Therefore polyclonal antibody obtained by this law has enantioselectivity to the antibody of relative medicine, wherein anti alpha RR-MET antibody
High sensitivity, specificity is good, can be used for the detection of relative medicine, remaining antibody can distinguish carbon chiral enantiomer.
Finally, it should be noted that the above embodiments are merely illustrative of the technical solutions of the present invention rather than protects to the present invention
The limitation of shield range can also be made on the basis of above description and thinking for those of ordinary skill in the art
Other various forms of variations or variation, there is no necessity and possibility to exhaust all the enbodiments.It is all of the invention
Made any modifications, equivalent replacements, and improvements etc., should be included in the protection of the claims in the present invention within spirit and principle
Within the scope of.
Claims (10)
1. a kind of isopropyl methoxalamine chirality haptens, which is characterized in that have the molecular structure as shown in formula (I):
2. the preparation method of isopropyl methoxalamine chirality haptens described in claim 1, which comprises the steps of:
S1. 3- mercaptopropionic acid, isopropyl methoxalamine are dissolved in ethyl alcohol, add KOH, the return stirring under the conditions of 65 DEG C~85 DEG C
5h~8h is reacted, isopropyl methoxalamine derivative is obtained；
S2. isopropyl methoxalamine derivative is carried out by chiral resolution using AD-H column, obtains four kinds of stereoisomer derivatives to get four
Kind isopropyl methoxalamine chirality haptens.
3. preparation method according to claim 2, which is characterized in that isopropyl methoxalamine described in step S1,3- mercaptopropionic acid,
The molal weight ratio of potassium hydroxide is 1:1:1~4.
4. isopropyl methoxalamine chirality haptens is in preparing isopropyl methoxalamine artificial antigen and/or antibody described in claim 1
Using.
5. a kind of isopropyl methoxalamine artificial antigen, which is characterized in that be the isopropyl methoxalamine chirality haptens as described in claim 1
It is obtained with carrier protein couplet.
6. isopropyl methoxalamine artificial antigen according to claim 5, which is characterized in that the carrier protein is bovine serum albumin
White or ovalbumin.
7. the preparation method of isopropyl methoxalamine artificial antigen described in claim 5, which comprises the steps of:
S1. isopropyl methoxalamine chirality haptens described in claim 1 is dissolved in dimethyl formamide solution at room temperature,
Isopropyl methoxalamine chirality haptens solution is obtained after mixing；
S2. carrier protein is dissolved in PBS solution and is uniformly mixed, obtain carrier protein solution；Again by isopropyl first grass obtained by step S1
Amine chirality haptens solution is added dropwise in carrier protein solution, obtains mixed solution, is stirred under the conditions of -4 DEG C, 100r/min mixed
It is even；
S3. carbodiimide is dissolved in dimethyl formamide solution, obtained Carbodiimide solution be slowly added dropwise while stirring to
In mixed solution obtained by step S2, in -4 DEG C of 8~12h of reaction；It is dialysed with phosphate buffer after reaction to get isopropyl is arrived
Alachlor artificial antigen.
8. preparation method according to claim 7, which is characterized in that the carrier protein and isopropyl methoxalamine chirality haptens
Molal weight ratio be 1:50~100, the volume fraction of dimethylformamide is 10~15%.
9. a kind of isopropyl methoxalamine polyclonal antibody, which is characterized in that be artificial by the isopropyl methoxalamine of claim 5 or 6
Antigen-immunized animal, then obtain animal blood serum and obtain after purification.
10. isopropyl methoxalamine polyclonal antibody described in claim 9 is in the kit of preparation detection isopropyl methoxalamine enantiomer
Application.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201910300552.7A CN110054576B (en) | 2019-04-15 | 2019-04-15 | Metolachlor chiral hapten, artificial antigen and antibody as well as preparation method and application thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201910300552.7A CN110054576B (en) | 2019-04-15 | 2019-04-15 | Metolachlor chiral hapten, artificial antigen and antibody as well as preparation method and application thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN110054576A true CN110054576A (en) | 2019-07-26 |
| CN110054576B CN110054576B (en) | 2021-03-12 |
Family
ID=67319041
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201910300552.7A Active CN110054576B (en) | 2019-04-15 | 2019-04-15 | Metolachlor chiral hapten, artificial antigen and antibody as well as preparation method and application thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN110054576B (en) |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112485430A (en) * | 2020-11-13 | 2021-03-12 | 北京勤邦生物技术有限公司 | Application of alachlor artificial antigen in enzyme linked immunosorbent assay kit |
| CN114316059A (en) * | 2021-12-31 | 2022-04-12 | 华南农业大学 | Variable region sequence of specific anti-metolachlor carbon chiral antibody and preparation of recombinant full-length antibody thereof |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103073634A (en) * | 2011-10-25 | 2013-05-01 | 天津科技大学 | Specific antibody against herbicide anilofos |
| CN106674138A (en) * | 2016-11-21 | 2017-05-17 | 河南科技大学 | Novel hapten for tebuconazole as well as synthetic method and application thereof |
-
2019
- 2019-04-15 CN CN201910300552.7A patent/CN110054576B/en active Active
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103073634A (en) * | 2011-10-25 | 2013-05-01 | 天津科技大学 | Specific antibody against herbicide anilofos |
| CN106674138A (en) * | 2016-11-21 | 2017-05-17 | 河南科技大学 | Novel hapten for tebuconazole as well as synthetic method and application thereof |
Non-Patent Citations (2)
| Title |
|---|
| 张炽坚等: "酶联免疫吸附法检测除草剂异丙甲草胺", 《食品科学》 * |
| 张璇等: "手性除草剂异丙甲草胺毒理研究进展", 《西北农林科技大学学报 自然科学版》 * |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112485430A (en) * | 2020-11-13 | 2021-03-12 | 北京勤邦生物技术有限公司 | Application of alachlor artificial antigen in enzyme linked immunosorbent assay kit |
| CN112485430B (en) * | 2020-11-13 | 2023-08-11 | 北京勤邦科技股份有限公司 | Application of alachlor artificial antigen in ELISA kit |
| CN114316059A (en) * | 2021-12-31 | 2022-04-12 | 华南农业大学 | Variable region sequence of specific anti-metolachlor carbon chiral antibody and preparation of recombinant full-length antibody thereof |
Also Published As
| Publication number | Publication date |
|---|---|
| CN110054576B (en) | 2021-03-12 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN1934133B (en) | Anti-influenza type A virus monoclonal antibody and immunoassay instrument using the antibody | |
| CN110054576A (en) | A kind of isopropyl methoxalamine chirality haptens, artificial antigen and antibody and its preparation method and application | |
| Liu et al. | Development of a monoclonal antibody based-ELISA for the detection of chloramphenicol in shrimp, feed and milk samples and validation by LC-MS/MS coupled with immunoaffinity clean-up | |
| CN101603965B (en) | Kit for quantitatively measuring PEG modified medicaments by ELISA competition method | |
| CN109265401A (en) | A kind of preparation method and application of iprodione haptens and antigen | |
| CN109307761A (en) | A kind of indirect competitive ELISA method detecting chaff propylhomoser | |
| CN109180519B (en) | Olaquindox metabolite antigen, antibody, enzyme-linked immunosorbent assay kit and detection method | |
| CN105624119B (en) | The general monoclonal antibody of anti-capsaicine, Dihydrocapsaicin, the synthetic capsaicin of hybridoma cell strain YQQD8 and its generation | |
| CN106496323A (en) | A kind of cluster selectivity ochratoxin artificial antigen and its preparation method of broad spectrum activity polyclonal antibody | |
| KR0161553B1 (en) | Immunological detection method of pyrethroid compound, haptenic compound, intermediate for the haptenic compound, imunogen and antibody | |
| CN109752531A (en) | The kit and its detection method of Fipronil in a kind of detection egg | |
| CN114276264B (en) | Fenfluramine hapten, artificial antigen, antibody, preparation method and application thereof | |
| CN100414296C (en) | Embedded high-pass three-dimensional biological detecting technique and agent box | |
| CN102627628A (en) | Chlorantraniliprole antigen, its preparation method and application | |
| CN102928407A (en) | Magnetic particle chemiluminescence kit for detecting avermectins, and applications thereof | |
| CN106380520B (en) | A kind of anti-Furaxone metabolite monoclonal antibody and its application | |
| CN108484752B (en) | Spiral shell worm ethyl ester antigen and the preparation method and application thereof | |
| CN110845619B (en) | Monoclonal antibody for identifying florfenicol, chloramphenicol and thiamphenicol and application thereof | |
| CN110117286B (en) | Heterocyclic amine 8-MeIQx hapten and antibody as well as preparation method and application thereof | |
| CN111675670A (en) | Citric acid-based dendritic hapten CAA, dendritic antigen and heavy chain antibody for directly detecting AMOZ and preparation method thereof | |
| CN102288761A (en) | Chloramphenicol testing kit and preparation method of same | |
| CN112415190A (en) | Time-resolved fluorescence immunochromatography method for detecting nicarbazin and application | |
| JP5690514B2 (en) | Azoxystrobin derivatives, antibodies to azoxystrobin or fragments thereof, and measurement kits and methods using these antibodies or fragments | |
| CN109917127A (en) | A kind of Spanon detection kit and its application | |
| CN117069580B (en) | Triclosan hapten as well as preparation method and application thereof |
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 |