酶联免疫法检测海洋贝类中的荧蒽
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摘要
分别以BSA和OVA为载体蛋白,利用活化酯法和混合酸酐法合成了荧蒽的免疫原和包被原,对新西兰大白兔进行免疫,制备出效价较高、特异性较好的荧蒽多克隆抗体。建立出一种检测贝类中荧蒽的间接竞争酶联免疫吸附方法(ic-ELISA),优化了包被原、一抗、二抗工作浓度,包被时间、抗原抗体反应时间及封闭液等实验检测条件。该方法特异性强,稳定性和灵敏度高,线性回归方程为y=19. 97x+15. 495 (R~2=0. 99),线性范围为1～500 ng/mL,IC_(50)为53 ng/mL,检出限为0. 94 ng/mL,回收率为81. 3%～102. 5%,RSD为2. 5%～8. 9%。将该法应用于实际贝类样品的检测,检测结果和HPLC相比,具有较好的相关性,该方法可用于实际样品中荧蒽的快速分析检测。
Fluoranthene immunogen and coating agent were synthesized by activated ester method and mixed anhydride method,with BSA and OVA as carrier proteins respectively. New Zealand white rabbits were immunized to obtain fluoranthene polyclonal antibody with higher titer and better specificity. An indirect competitive enzyme-linked immunosorbent assay( ic-ELISA) for the detection of fluoranthene in shellfish was established,and the working conditions was optimized. This method has high specificity,stability and sensitivity. The linear regression equation is y = 19. 97 x +15. 495( R~2= 0. 99),with the linear range of 1-500 ng/mL. The IC_(50) is 53 ng/mL,the limit of detection is 0. 94 ng/m L,the recoveries are 81. 3 %-102. 5 %,and the RSDs are 2. 5 %-8. 9%. This method was applied to the detection of fluoranthene in shellfish samples. The results had good correlation with HPLC,which showed this method can be used for rapid detection of fluoranthene in real samples.
Fluoranthene immunogen and coating agent were synthesized by activated ester method and mixed anhydride method,with BSA and OVA as carrier proteins respectively. New Zealand white rabbits were immunized to obtain fluoranthene polyclonal antibody with higher titer and better specificity. An indirect competitive enzyme-linked immunosorbent assay( ic-ELISA) for the detection of fluoranthene in shellfish was established,and the working conditions was optimized. This method has high specificity,stability and sensitivity. The linear regression equation is y = 19. 97 x +15. 495( R~2= 0. 99),with the linear range of 1-500 ng/mL. The IC_(50) is 53 ng/mL,the limit of detection is 0. 94 ng/m L,the recoveries are 81. 3 %-102. 5 %,and the RSDs are 2. 5 %-8. 9%. This method was applied to the detection of fluoranthene in shellfish samples. The results had good correlation with HPLC,which showed this method can be used for rapid detection of fluoranthene in real samples.
引文
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