Kim Hee-Joo, González-Techera Andrés, González-Sapienza Gualberto G, Ahn Ki Chang, Gee Shirley J, Hammock Bruce D
Department ofEntomology and UCD Cancer Research Center, University of California, Davis, California 95616, USA.
Environ Sci Technol. 2008 Mar 15;42(6):2047-53. doi: 10.1021/es702219a.
Competitive immunoassays for the detection of small analytes, such as pesticides and their metabolites, use haptens that compete with the target compounds for binding to the antibody. This competing hapten can be either the same as the immunizing hapten (homologous assay) or structurally modified mimics of the immunizing hapten (heterologous assay). Polyclonal antibody-based heterologous immunoassays have shown superior sensitivities to homologous ones, butthe synthesis of heterologous haptens may be time-consuming, requiring expertise in synthetic chemistry. In this work we demonstrate that phage display peptide libraries can be used as a source of phage-borne peptidomimetics to facilitate the development of sensitive heterologous assays. Different strategies for the isolation of these peptides were explored using two metabolites of pyrethroid insecticides. The sensitivities of the best competitive phage heterologous enzyme-linked immunosorbent assays were 13 fold and 100 fold better than the homologous assay, for the glycine conjugate of trans-3-(2,2-dichlorovinyl)-2,2-dimethylcyclopropane-1-carboxylic acid and 3-phenoxybenzoic acid, respectively. The phage particles were highly versatile as tracer reagents, allowing the use of enzymatic, chemiluminescent, or immuno-polymerase chain reaction detection. The data presented here shows a new systematic procedure that enables the fast generation of several competing haptens for the rapid development of sensitive heterologous immunoassays.
用于检测小分子分析物（如农药及其代谢物）的竞争性免疫分析，使用与目标化合物竞争结合抗体的半抗原。这种竞争性半抗原可以与免疫原性半抗原相同（同源分析），也可以是免疫原性半抗原的结构修饰模拟物（异源分析）。基于多克隆抗体的异源免疫分析已显示出比同源分析更高的灵敏度，但异源半抗原的合成可能耗时，需要合成化学方面的专业知识。在这项工作中，我们证明噬菌体展示肽库可作为噬菌体携带的拟肽来源，以促进灵敏异源分析的开发。使用拟除虫菊酯类杀虫剂的两种代谢物探索了分离这些肽的不同策略。对于反式-3-(2,2-二氯乙烯基)-2,2-二甲基环丙烷-1-羧酸和3-苯氧基苯甲酸的甘氨酸共轭物，最佳竞争性噬菌体异源酶联免疫吸附分析的灵敏度分别比同源分析高13倍和100倍。噬菌体颗粒作为示踪剂具有高度通用性，允许使用酶促、化学发光或免疫聚合酶链反应检测。此处呈现的数据显示了一种新的系统程序，该程序能够快速生成几种竞争性半抗原，以快速开发灵敏的异源免疫分析。
