溴氰菊酯酶联免疫吸附分析和胶体金免疫层析分析方法的建立  

Enzyme Linked Immunosorbent Assay and Gold Nanoparticle-based Lateral Flow Immunoassay for Rapid Detection of Deltamethrin

作　　者：崔盼盼 王贵绣 吴艳玲 张召贤 丁园 华修德[1] CUI Pan-Pan;WANG Gui-Xiu;WU Yan-Ling;ZHANG Zhao-Xian;DING Yuan;HUA Xiu-De(College of Plant Protection,Nanjing Agricultural University,Nanjing 210095,China;College of Resources and Environment,Anhui Agricultural University,Hefei 230036,China)

机构地区：[1]南京农业大学植物保护学院,南京210095 [2]安徽农业大学资源与环境学院,合肥230036

出　　处：《分析化学》2024年第6期828-837,共10页Chinese Journal of Analytical Chemistry

基　　金：国家自然科学基金项目(No.32102258);安徽省重点研究与开发计划项目(No.2022i01020009)资助。

摘　　要：制备了9株分泌溴氰菊酯单克隆抗体的杂交瘤细胞株,选取灵敏度最高的单克隆抗体4D4E11建立了间接竞争酶联免疫吸附分析方法(ic-ELISA)和胶体金免疫层析分析方法(LFIA)用于检测溴氰菊酯。ic-ELISA的最优工作溶液为0.01 mol/L磷酸盐缓冲溶液(PB,NaH_(2)PO_(4)/Na_(2)HPO_(4),pH 7.4),含20%甲醇和0.2 mol/L Na Cl;LFIA的最优工作溶液为0.01 mol/L PB缓冲溶液(pH 7.4),含5‰Tween-20、10%甲醇和1 mol/L NaCl。在最优条件下,ic-ELISA对溴氰菊酯的半数抑制浓度(IC_(50))为10.60 ng/mL,检出限(IC10)为1.43 ng/mL;LFIA对溴氰菊酯的可视检出限为0.5μg/m L。特异性评价实验结果表明,ic-ELISA和LFIA对溴氰菊酯的8种结构类似物无明显交叉反应,特异性强。采用Ic-ELISA检测番茄、结球甘蓝和油麦菜添加样品的平均回收率为79.8%~92.6%,相对标准偏差为0.8%~5.5%;采用LFIA检测番茄、油麦菜和结球甘蓝添加样品的结果与添加浓度相符。此外,ic-ELISA和LFIA对油麦菜盲样中溴氰菊酯的检测结果与高效液相色谱的检测结果一致。实验结果表明,建立的ic-ELISA和LFIA灵敏度高、准确性好,为农产品中溴氰菊酯残留的快速检测提供了新方法。In this study,nine hybridoma cells secreting monoclonal antibodies against deltamethrin were prepared,and the monoclonal antibody 4D4E11 with best sensitivity was selected to develop indirect enzymelinked immunosorbent assay(ic-ELISA)and gold nanoparticle-based lateral flow immunoassay(LFIA)for detection of deltamethrin.The optimal working buffer for ic-ELISA was 0.01 mol/L phosphate buffer(pH 7.4)containing 0.2 mol/L NaCl and 20%methanol,while 0.01 mol/L phosphate buffer(pH 7.4)containing 1 mol/L NaCl,5‰Tween-20 and 10%methanol for LFIA.Under the optimal conditions,the half inhibition concentration(IC_(50))and limit of detection(IC10)of ic-ELISA were 10.60 ng/mL and 1.43 ng/mL respectively,and the limit of detection of the developed LFIA was 0.5μg/mL.The developed ic-ELISA and LFIA showed no cross-reactivities(CRs)with eight kinds of analogues of deltamethrin,which indicated the excellent specificity of proposed immunoassays.The average recoveries of the ic-ELISA in spiked tomato,cabbage and lettuce samples were 79.8%–92.6%with relative standard deviations of 0.8%–5.5%.The detection results of LFIA were consistent with the spiked concentrations in the range of 1–5 mg/kg.Meanwhile,the results of ic-ELISA and LFIA showed close correlation with high performance liquid chromatography(HPLC)in the test of blind lettuce samples.The experimental results demonstrated that the two immunoassays proposed here were suitable for rapid detection of deltamethrin with high sensitivity and high accuracy.

关 键 词：溴氰菊酯 单克隆抗体 间接竞争酶联免疫吸附法 胶体金免疫层析法 

分 类 号：TS255.7[轻工技术与工程—农产品加工及贮藏工程]