海水中邻苯二甲酸二乙酯的快速检测方法建立

Establishment of rapid detection method for diethyl phthalate in sea water

为构建海水中邻苯二甲酸二乙酯(DEP)快速检测方法,采用胶体金酶联免疫层析技术,利用抗DEP单克隆抗体杂交瘤细胞株进行腹水制备。以邻苯二甲酸单乙酯(MEP)-OVA为包被抗原,羊抗鼠IgG为二抗,优化抗原抗体配对组合和胶体金-抗体标记条件,最后组装获得快速检测DEP胶体金免疫层析试纸条。利用包括DEP在内的12种邻苯二甲酸酯类化合物标准溶液检测试纸条的灵敏度和特异性,并利用人造海水作为稀释溶剂检测试纸条的海水环境适应性。结果显示:纯化后抗体效价最高可以达到1∶30 000以上,胶体金溶液与抗体最佳标记条件为pH=7.2,抗体最佳标记量为6μL·mL^(-1);所制备胶体金酶联免疫层析(GICA)试纸条对DEP的检测特异性较高,在人造海水条件下试纸条对DEP的检测限可达20μg·L^(-1),适用于海水中DEP的快速检测。

A rapid detection method for DEP in sea water was developed using colloidal gold enzyme-linked immunochromatographic technique coupled with the highly efficient and stable hybridoma cell line secreting anti-DEP McAb(prepared by our research group) to produce ascites. When MEP-OVA and goat IgG anti-mouse were used as the coating antigen and the secondary antibody, respectively, the antigen-antibody pairing combination and colloidal gold-antibody labeling conditions were optimized, and the colloidal gold enzymelinked immunochromatographic(GICA) strips for the rapid DEP detection were prepared. The sensitivity and specificity of the strips were tested using twelve standard solutions of PAEs including DEP. The effect of sea water on the performance of the strips was detected by using artificial seawater as a dilution solvent. The results showed that the antibody titer is up to 1∶30 000. The optimal labeling pH for colloidal gold solution and antibody was 7.2. The antibody labeling amount is 6 μL·mL-1.A high specificity of DEP detectionwas identified forthe GICA strip with 20 μg·L-1 DEP detection limit in artificial seawater.This indicates that the GICA strip is suitable for rapid detection of DEP in sea water.