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A sensitive immunoassay based on direct hapten coated format and biotin-streptavidin system for the detection of chloramphenicol
Authors:Na Sai  Yiping Chen  Nan Liu  Pu Su  Zhijiang Zhou  Huanying Zhou  Bao an Ning
Affiliation:a School of Chemical Engineering and Technology, Tian Jin University, Tianjin 300072, China
b College of Food Science and Technology, Huazhong Agricultural University, Wuhan 430070, China
c Tianjin Key Laboratory of Risk Assessment and Control Technology for Environment & Food Safety; Institute of Hygiene and Environmental Medicine, Tianjin 300050, China
d College of Bioscience and Biotechnology, YangZhou University, JiangSu 225009, China
Abstract:A novel enzyme liked immunosorbent assay (ELISA) was developed for the detection of chloramphenicol (CAP). In this assay, the small molecular hapten (Hap) was directly coated on the surface of microtiter plates and biotin-streptavidin system (BSAS) was employed to improve the sensitivity of immunoassay (BSAS-direct Hap coated ELISA). The surface of microtiter plates was treated with glutaraldehyde (GA) polymer network to introduce aldehyde group, which was used to cross-link with amino group of CAP. Compared with conventional ELISA (the plates were coated with Hap-carrier protein conjugates), the modified plates presented significantly high antibody and antigen (Ab-Ag) affinity and showed excellent stability. And then the biotinylated monoclonal antibody (mAb) and HRP-labeled streptavidin were employed in this assay for amplification of signals. The sensitivity of BSAS-direct Hap coated ELISA was increased by approximately 20-folds and the stability was also improved greatly compared to conventional ELISA. Its 50% inhibition concentration (IC50) for CAP was 10.5 ng mL−1 and the limit of detection (LOD) was 0.2 ng mL−1 after optimization of reaction conditions. To our knowledge, this was one of the most sensitive immunoassay for CAP yet reported. In sample analysis, the results of CAP detected by this assay were in accordance with which obtained by conventional ELISA and high performance liquid chromatography (HPLC). Therefore, it is an attractive alternative compared to conventional immunoassays in routine supervision for residue detection in food and environment.
Keywords:Polymerized glutaraldehyde   Biotin-streptavidin system   Immunoassay   Chloramphenicol
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