Development of indirect competitive fluorescence immunoassay for 2,2′,4,4′-tetrabromodiphenyl ether using DNA/dye conjugate as antibody multiple labels

作     者：Zi-Yan Fan Young Soo Keum Qing-Xiao Li Weilin L.Shelver Liang-Hong Guo 

作者机构：State Key Laboratory of Environmental Chemistry and EcotoxicologyResearch Center for Eco-environment SciencesChinese Academy of SciencesBeijing 100085China College of Life and Environmental SciencesKonkuk University in Korea1 Hwayang-dongGwangjin-guSeoul 143-701Korea Department of Molecular Bioscience and BioengineeringUniversity of Hawaii at Manoa1955 East-West RoadHonoluluHI 96822USA USDA-ARS Bioscience Research Laboratory1605 Albrechht BoulevardFargoND 58105USA 

出 版 物：《Journal of Environmental Sciences》 (环境科学学报（英文版）)

年 卷 期：2012年第24卷第7期

页      面：1334-1340页

核心收录：

学科分类：0710[理学-生物学] 071010[理学-生物化学与分子生物学] 081704[工学-应用化学] 07[理学] 08[工学] 0817[工学-化学工程与技术] 070302[理学-分析化学] 0703[理学-化学] 

基　　金：supported by the Chinese Academy of Sciences (No. KSSCX2-YW-G-059) the National Hi-Tech Research and Development Program of China (No.2007AA06A407) the National Natural Science Foundation of China (No. 20825519, 20890112, 20921063) 

主　　题：polybrominated diphenyl ethers fluorescence immunoassay microplate multiple labeling 

摘      要：An indirect competitive fluorescence immunoassay using a DNA/dye conjugate as antibody multiple labels was developed on 96-well plates for the identification and quantification of 2,2＇,4,4＇-tetrabromodiphenyl ether （BDE-47） in aqueous samples. A hapten, 2,4,2＇- tribromodiphenyl ether-4＇-aldehyde, was synthesized, and was conjugated to bovine serum albumin to form a coating antigen. Specific recognition of the antigen by anti-PBDE antiserum was confirmed by a surface plasmon resonance measurement. In the immunoassay, the coating antigen was adsorbed on a 96-well plate first, and a sample, antiserum and biotinylated goat anti-rabbit secondary antibody were then added and reacted sequentially. A biotinylated, double-stranded DNA with 219 base pairs was attached to the secondary antibody by using streptavidin as a molecular bridge. In situ multiple labeling of the antibody was accomplished after addition of a DNA-binding fluorescent dye, SYBR Green I. The working range of the immunoassay for the BDE-47 standard was 3.1-390 ~tg/L, with an IC50 value of 15.6 Ixg/L. The calculated LOD of the immunoassay is 0.73 Ixg/L. The immunoassay demonstrated relatively high selectivity for BDE-47, showing very low cross-reactivity （〈 3%） with BDE-15, BDE-153 and BDE-209. With a spiked river water sample containing 50 Izg/L BDE-47, quantification by the immunoassay was 41.9 ~tg/L, which compared well with the standard GC-ECD method （45.7 Ixg/L）. The developed immunoassay provides a rapid screening tool for polybrominated diphenyl ethers in environmental samples.