双酚A残留酶联免疫分析方法的研究及应用  

Study and Application of an Enzyme-linked Immunosorbent Assay for Detection of Bisphenol A Residues

作　　者：张丽莎[1] 吕型超 陈涵倩 邓杰[1] 潘熙萍[1] 施曼玲[1] 

机构地区：[1]杭州师范大学生命与环境科学学院,浙江杭州310036

出　　处：《核农学报》2014年第2期234-239,共6页Journal of Nuclear Agricultural Sciences

基　　金：浙江省科技厅公益技术研究农业项目(2011C22100);浙江省环境保护厅科研计划项目(2013B012)

摘　　要：为了建立双酚A残留的酶联免疫分析方法,采用碳化二亚胺法将半抗原双酚酸(BHPVA)分别与卵清蛋白(OVA)和牛血清蛋白(BSA)偶联,制备免疫原BHPVA-OVA和包被原BHPVA-BSA。基质辅助激光解析电离飞行时间质谱(MALDI-TOF-MS)检测表明BHPVA与BSA和OVA的偶联比分别为12∶1和16∶1。BHPVA-OVA免疫新西兰大白兔获得抗双酚A的多克隆抗体,抗体ELISA效价为1∶1×107。基于该抗体建立双酚A间接竞争ELISA(ciELISA)检测方法,其检测的线性范围为0.02-100ng·mL-1,线性回归方程y=9.0326 ln x+39.105,R2=0.9975,双酚A的检测限为IC10=0.04ng·mL-1。采用所建立的ciELISA方法均检测出8种塑料食品袋中的双酚A残留,迁移析出量分别从4.9到31mg·L-1,批内和批间变异系数均小于12.48%,说明本试验所建立ciELISA法可有效地用于塑料食品袋中的双酚A迁移量的测定,不仅具有高度的灵敏性,而且具有较好的稳定性。In order to establish an enzyme-linked immunosorbent assay （ELISA） method for detecting bisphenol A （BPA） residues in food plastic bags, diimine carbonization methods were used to synthesize an immunogen （BHPVA- OVA） and a coating antigen （BHPVA-BSA） by conjugating a hapten, 4,4 -Bis（4 -hydroxyphenyl） valeric acid （BHPVA） to ovalbumin （OVA） or bovine serum albumin （BSA） respectively. The results of matrix-assisted laser desorption ionization-time of flight mass spectrometry （MALDI-TOF-MS） showed that the ratios of hapten to OVA and BSA for hapten conjugation were 16：1 and 12：1. New Zealand white rabbits were immunized with BHPVA-OVA, and polyclonal antibodies against BPA were raised with ELISA titers of 1 ： 1 x 107. The antibodies were used in an indirect competitive ELISA （ciELISA） for BPA detection and this assay has a detection limit （ICL0） of 0.04ng. mL-~ with a linear working range of 0.02 to 100ng. mL-1. The regression equation was y = 9. 0326 In x ＋ 39. 105,R2 = 0. 9975. BPA residues were found in all eight types of food plastic bags tested by ciELISA, and these ranged from 4.9 to 31mg- L-1 with the intra-and inter-assay CV below 12.48 %. These results indicate that the ciELISA assay can be used effectively to detect BPA contamination in food plastic bags, and the method has not only high sensitivity but also good reliability.

关 键 词：双酚A 多克隆抗体 间接竞争酶联免疫吸附法 塑料食品袋 

分 类 号：TS206[轻工技术与工程—食品科学]