快速定量分析牛乳中α-乳白蛋白的竞争酶联免疫吸附法

Competitive enzyme-linked immunosorbent assay for rapid quantitative analysis of α-lactalbumin in bovine milk

建立一种灵敏度高和特异性好的直接竞争ELISA法来定性和定量检测牛乳中α-乳白蛋白（α-LA）.通过使用牛乳α-LA多次免疫新西兰大白兔制备多克隆抗体,并采用过碘酸钠氧化法制备辣根过氧化物酶（HRP）标记的α-LA,建立检测α-LA的直接竞争ELISA法.所制备的多克隆抗体特异性较高,无明显交叉反应,效价为1∶80 920.酶标抗原的标记率达66.67%.直接竞争ELISA法标准曲线相关系数为0.998 57,回收率在86.00%-117.50%之间,重复性高,变异系数〈5%.与国标电泳方法进行样品检测比较,结果显示该方法可有效地用于食品中α-LA的快速定性和定量检测.

A sensitive and specific competitive enzyme-linked immunosorbent assay （cELISA） for qualitative and quantitative analysis of α-laetalbumin （α-LA） is described. New Zealand white rabbits were immunised to obtain α-LA PcAb. By using sodium periodate oxidative method, the α-LA labeled by horseradish peroxidase （HRP） was prepared. Then cELISA was developed for α-LA quantitative analyses. The prepared PcAb has high specificity without obvious cross-reactivity and its titer is 1 ： 80 920. The labeling rate of the enzyme-labeled antigen is 66.67%. The correlation coefficient of the established cELISA was 0. 998 57, and the recoveries were estimated to be in the range from 86.00% to 117.50%. The variation coefficient is less than 5%. Compared with the national standard electrophoresis method,the established eELISA was useful analytical tool for rapid qualitative and quantitative analysis of α-LA in food samples.