基于高特异性单克隆抗体的间接竞争ELISA检测食品中胭脂红的研究

朱琳; 陈子键; 罗林; 王宇; 钟玉心; 沈玉栋; 孙远明; 徐振林

Study on Detection of Carmine by Indirect Competitive ELISA Based on Highly Specific Monoclonal Antibody

DOI：

KeyWord:carmine hapten design monoclonal antibody indirect competitive enzyme-linked immunosorbent assay （icELISA） food

Author	Institution
ZHU Lin,CHEN Zi-jian,LUO Lin,WANG Yu,ZHONG Yu-xin,SHEN Yu-dong,SUN Yuan-ming,XU Zhen-lin	1. Guangdong Provincial Key Laboratory of Food Quality and Safety，College of Food Science，South China Agricultural University，Guangzhou ，China; 2. Guangzhou Institute for Food Control， Guangzhou ，China

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Abstract:

Carmine is one of the most widely used synthetic pigments in foods．However，a certain dose of carmine may result in teratogenicity，sensitization，neurotoxicity，reproductive and developmental toxicity．In order to develop a simple，rapid and accurate analytical method for screening carmine residues in food，a novel hapten against carmine was designed and synthesized in this paper. It was conjugated to lactoferrin by diazotization method to obtain immunogen and bovine serum albumin by glutaraldehyde method to obtain coating antigen．After immunizing the Bal b/c mice by immunogen，the spleen cells were fused with SP 2/0 to obtain the hybridoma screening monoclonal antibodies with high specificity to carmine．Negligible cross-reactivity with carmine analogous such as amaranth，lemon yellow，brilliant blue and sunset yellow was obtained．Based on the obtained monoclonal antibody，a simple，rapid and sensitive indirect competitive enzyme-linked immunosorbent assay（icELISA） was developed and validated. After optimizing the working conditions，the proposed icELISA showed a good sensitivity for carmine detection， with a half inhibition rate（IC50） of 10.1 ng/mL，a limit of detection（IC10） of 0.98 ng/mL and a linear range of 2－50 ng/mL．The proposed assay was then applied to detection of carmine in sprite and hawthorn strips samples．Recoveries of 93.0%－100% for sprite samples and 93.0%－113% for hawthorn strips samples were obtained with relative standard deviations of 3.2%－3.9% and 9.7%－12%，respectively，which were validated by standard high performance liquid chromatography coupled with ultraviolet-visible spectroscopy（HPLC－UV）．Results showed that the proposed icELISA was an ideal tool for the rapid screening of carmine residues in food samples.