间接竞争化学发光酶联免疫分析方法检测禽肉中金刚烷胺和氯霉素残留

Determination of Amantadine and Chloramphenicol Residues in Poultry Meat by Indirect Competitive Chemiluminescence Enzyme-Linked Immunosorbent Assay

为快速检测禽肉中残留的金刚烷胺和氯霉素,结合高效的前处理方法,建立检测金刚烷胺和氯霉素的间接竞争化学发光酶联免疫分析方法(indirect competitive chemiluminescence enzyme-linked immunosorbent assay,ic-CLEIA)。对包被原与抗体的最佳稀释倍数、竞争时间、酶标二抗稀释倍数进行优化。结果表明:金刚烷胺的IC50为0.33μg/L,线性范围为0.06~1.77μg/L;氯霉素的IC50为0.039μg/L,线性范围为0.010~0.179μg/L。样品经乙酸乙酯和碳酸钾溶液提取后,取上清液用氮气吹干,净化后进行检测。金刚烷胺和氯霉素的加标回收率分别为92.55%~105.14%和92.00%~112.50%,变异系数均低于10.48%,且实际样品检测结果与仪器方法相关性良好(R2>0.98),表明建立的ic-CLEIA方法具有良好的准确度和可靠性。该方法将为涉及多种药物残留的免疫快速检测方法及相应的前处理技术提供技术依据。

An indirect competitive chemiluminescent enzyme-linked immunosorbent assay(ic-CLEIA)was developed for the rapid detection of amantadine and chloramphenicol residues in poultry meat based on efficient sample pretreatment.The dilution factors of coating antigen,antibody and horseradish peroxidase(HRP)-labeled secondary antibody,and competition time were optimized.The results showed that the half-maximum inhibition concentration(IC50)and the linear range were 0.33μg/L and 0.06–1.77μg/L for amantadine,and 0.039μg/L and 0.010–0.179μg/L for CAP,respectively.Samples were sequentially extracted with ethyl acetate and potassium carbonate solution,and then the extract was blown to dryness with nitrogen gas and purified by n-hexane extraction before being analyzed.The recoveries of amantadine and chloramphenicol from spiked samples were in the ranges of 92.55%–105.14%and 92.00%–112.50%,respectively,and the coefficients of variation were both less than 10.48%.Furthermore,the results determined by ic-CLEIA were highly correlated with those of instrumental analysis method(R2>0.98),which demonstrates that the established ic-CLEIA method has high accuracy and good reliability.This method represents a rapid immunoassay for the detection of veterinary drug residues in poultry meat.