Abstract
The aim of this study was to prepare B-group aflatoxins(BGAFs) antibody with strong specificity and good eurytopicity. According to the molecular structure and active site of aflatoxin B1 (AFB1), the BGAFs artificial antigen AFB1-BSA was prepared by 6 methods such as oxime active ester(OAE)，methylation of ammonia(MOA)，mixed anhydride(MA)，semi acetal(SA)，epoxide(EP) and enol ether derivative(EED) and identified by UV and SDS-PAGE. Polyclonal antibodies against AFB1(AFB1 pAb) were prepared by immunizing New Zealand rabbits with AFB1-BSA, and the titers of AFB1 pAb was detected by indirect ELISA, the sensitivity of AFB1 pAb was analyzed by indirect competitive ELISA(icELISA) and the specificity and eurytopicity of AFB1 pAb was analyzed by cross-reactivity(CR) test. The results showed that AFB1-BSA was synthesized successfully and the best one was OAE method among 6 synthesis methods of BGAFs artificial antigen and its conjugation ratio of AFB1 to BSA was about 8.46∶1. The immune efficacy of OAE method was the best, its AFB1 pAb had high titers of 1∶（1.28×104） by indirect ELISA, a good sensitivity with the 50% inhibition concentration(IC50) of 10.32 μg/L to AFB1 by icELISA and a high CR to AFB2 of 75.21%, AFG1 of 44.13%, AFG2 of 14.72%, AFM1 of 16.36% and AFM2 of 1.44%, respectively. In this study, AFB1 pAbs with high titer, sensitivity, specificity and eurytopicity were prepared, which laid a matter and technical foundation for the establishment of BGAFs immunoassay.
Highlights
IntroductionImmunoassay has become a techondary metabolites containing similar molecular structures nology because of its strong specificity, high sensitivity, simple (difuran ring and oxyheteronaphthalidone)
Aflatoxins (AFs) are a group of toxic sec- immunoassay
The results showed that aflatoxin B1 (AFB1)-BSA was synthesized successfully and the best one was OAE method among 6 synthesis methods of BGAFs artificial antigen and its conjugation ratio of AFB1 to BSA was about 8.46∶1
Summary
Immunoassay has become a techondary metabolites containing similar molecular structures nology because of its strong specificity, high sensitivity, simple (difuran ring and oxyheteronaphthalidone). They are produced operation, large-scale screening and on-site detection. BGAFs include AFB1 and design of different hapten molecules, antigen synthesis and AFB2, both of which are closely related to food pollution, and comparative analysis of antibody characteristics. In this study, both exist at the same time, mainly AFB1, AFB2 pollution is AFB1 was used as the starting material for the reaction. There are two regulations for the max- hapten molecular design and antigen synthesis methods, and imum residue limits (MRLs) of BGAFs in foods. the current their characteristics were analyzed to screen out the best hap-
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