糙米、土壤和田水中三唑磷残留的化学发光酶免疫分析方法

Development of chemiluminescent enzyme immunoassay for the determination of triazophos residues in rice grain,soil and paddy water

建立三唑磷间接竞争化学发光酶免疫法（indirect competitive chemiluminescent enzyme immunoassay，icCLEIA）．以鲁米诺和过氧化氢作为化学发光底物，通过对工作浓度、化学发光增敏液、反应缓冲液、有机助溶剂与浓度等条件的优化，确立方法的最佳工作条件．该方法对三唑磷检测线性范围为0．097～12．5ng/mL，ICso为1．23ng/mL，检出限可达0．14ng/mL．将优化的方法应用到稻田水样、土样、糙米样品中三唑磷残留检测，样品的添加回收率为67％～113％，变异系数为0．08％～24．53％．研究表明该方法可用于上述样品中三唑磷残留的快速检测，并可为三唑磷化学发光试剂盒的开发应用提供依据．

Triazophos is now widely used in agriculture. Its residues may cause health implications for living organisms including humans. There has been a greater demand for developing analytical methods with very high sensitivity. Immunochemical techniques provide a simple and economical alternative to traditional methods in order to meet such demands for determination of triazophos residues. The aim of this work was to develop an indirect competitive chemiluminescent enzyme immunoassay （icCLEIA） with an anti-mouse secondary antibody conjugated to peroxidase for the residual determination of triazophos. Luminol solution was used as the substrate of horseradish peroxidase. First, the optimal coating conjugate concentration and anti-triazophos monoclonal antibodies dilution were studied. Then the chemiluminescence substrate solution was obtained after a variety of conditions were optimized, including the selection of buffer and enhancer, concentrations of luminol, p-iodophenol and HzOz. On the basis of it, the effects of several physicochemical parameters on icCLEIA performance were studied and optimized, such as the effects of ionic strength and pH, and the tolerance to organic solvents （methanol, acetonitrile, acetone）. After the condition optimization, the standard curve of CLEIA competitive reaction for triazophos was established. Afterwards, theoptimized method was applied to the detection of triazophos residues in paddy water, soil, and rice grain samples, and the precision and accuracy were analyzed as well.