摘要
建立了测定水产品中庆大霉素(GM)残留的直接竞争酶联免疫吸附测定法(dcELISA),为进一步开发商业化的ELISA试剂盒提供技术支持。通过碳二亚胺法和戊二醛法分别制备了免疫原GM-KLH和包被原GM-OVA,经动物免疫后获取的多克隆抗体与辣根过氧化物酶偶联制备酶标抗体GM-PAb-HRP。优化确立了dcELISA最佳检测条件:包被原质量浓度0.2μg/m L,GM-PAb-HRP稀释度1/3 200,抗体反应时间40 min,缓冲液为PBS(pH 8.0,10 mmol/L)。该方法的半抑制浓度(IC_(50))为0.99μg/L,定量检测线性范围为0.27~3.64μg/L。水产样品加标回收率为77.7%~104.5%,RSD为6.7%~13.2%,最低检测限为2.0μg/kg。该方法可用于水产品中庆大霉素残留的快速测定,为食品安全风险监测提供一种有效的技术手段。
A direct competitive enzyme -linked immunosorbent assay （dcELISA） was developed for rapid determina- tion of gentamiein in aquatic product, which exhibited the potential to develop commercial ELISA kits. The immuno- gen （ GM - KLH） and coating antigen （ GM - OVA） were synthesized by the method of earbodiimide and glutaralde- hyde, respectively. Then, the polyelonal antibody obtained was coupled to horseradish peroxidase for enzyme -la- beled antibody （GM- PAb- HRP）. The optimized deELISA conditions were as follows： the dilution ratio of GM - PAb- HRP, 1/3 200; coating antigen concentration, 0.2 μg/mL; reaction time of antibody, 40 min; dilution solu- tion, PBS （pH 8.0, 10 mmol/L）. The half inhibition concentration （ICs0） was 0.99 μg/L and the linear range （IC20 - IC80） was 0.27 - 3.64 μg/L. Recoveries from spiked aquatic products were in the range of 77.7% 104.5%, with relative standard deviation ranging from 6.7% to 13.2%. The limit of detection （LOD） of the dcELISA for gentamiein in aquatic products was 2.0 μg/kg. The developed method was suitable for rapid determina- tion of gentamiein residues in aquatic product, which would be a useful tool in regular risk monitoring program for food safety.
出处
《中国渔业质量与标准》
2017年第2期36-42,共7页
Chinese Fishery Quality and Standards
基金
国家农产品质量安全风险评估重大专项(GJFP201501003)
广东省水产品质量安全专项(GDSCZA2015009)
中国水产科学研究院南海水产研究所基本科研业务费(2015TS17,2015TS18)
