新霉素的直接竞争酶联免疫分析法.pdf

212 2011, Vol. 32, No. 10 食品科学 ※分析检测 徐乃丰，胥传来，匡 华，屈昌龙，许 阳，彭池方 * (江南大学食品学院，江苏 无锡 214122) 首先用碳二亚胺(EDC)法将新霉素(NEO)偶联于载体蛋白- 卵清白蛋白(ovalbumin，OVA)，合成包被原OVA- NEO ，SDS进行鉴定；用高碘酸钠法连接辣根过氧化物酶(horseradish peroxidase ，HRP) ，制备酶标抗原 NEO-HRP 并建立直接 ELISA 检测方法。通过一系列参数的优化，包括包被溶液、封闭溶液、竞争时间、抗体 稀释液、pH 值、反应温度、显色时间等，最终得到其 IC20 (抑制率为 20% 时的标准溶液质量浓度) ＜ 1ng/mL、半 数抑制量(IC 2 50)为 7.6ng/mL ，线性方程为y = －0.2798x ＋0.7456 ，R =0.991 。直接ELISA 总耗时只需大约 1h。 新霉素；间接竞争酶联免疫吸附试验；直接竞争酶联免疫吸附试验 Direct Competitive Enzyme-linked Immunosorbent Assay (ELISA) for Neomycin XU Nai-feng ，XU Chuan-lai ，KUANG Hua ，QU Chang-long ，XU Yang ，PENG Chi-fang* (School of Food Science and Technology, Jiangnan University, Wuxi 214122, China) Abstract ：The neomycin (NEO) was coupled with ovalbumin (OVA) to form a coating antigen NEO-OVA by EDC method and determined by SDS. The NEO was coupled with horseradish peroxidase (HRP) by NaIO4 method to establish enzymatic tracer NEO-HRP and direct competitive ELISA. Then, the parameters coating liquid, pH, incubation time and incubation temperature of direct ELISA were optimized. The IC20 value (20% inhibitory concentration) was less than 1 ng/mL 2 and IC50 value (50% inhibitory concentration) was 7.6 ng/mL. The linear equation was y = －0.2798x ＋0.7456, R = 0.991. The total consumption time of direct competitive ELISA was 1 h. Key words ：neomycin ；indirect competitive enzyme-linked immunosorbent assay ；direct competitive enzyme-linked immunosorbent assay 中图分类号：S859.84 文献标识码：A 文章编号：1002-6630(2011)10-0212-06 动物性食品的安全问题已成为社会各界极为关注的 1 材料与方法 食品安全