农畜产品中三聚氰胺间接竞争ELISA检测方法的建立

利用制备的抗三聚氰胺单克隆抗体建立了间接竞争ELISA方法,采用棋盘滴定法确定最佳三聚氰胺-OVA包被浓度为1.0 μg/mL,单抗最佳稀释度为1∶1 000,最佳的封闭液为50 g/L的脱脂乳,标准曲线的线性方程为y=-15.12x+107.3(R2=0.997).批内变异系数(CV)为1.07％～5.53％,批间CV值为3.33％～11.92％,确定此方法精确度良好.利用建立的间接竞争ELISA方法,测定添加有三聚氰胺标准溶液的样品(宠物食品、小麦面筋、饲料、牛乳、鸡蛋),得到三聚氰胺在各样品中的回收率范围分别为宠物食品73.8％～84.32％,小麦面筋67.3％～88.27％,饲料70.3％～85.9％,牛乳83.47％～89.76％,鸡蛋76.2％～87.29％.确定可以利用此间接竞争ELISA方法检测样品中的三聚氰胺含量.

Development of an Indirect Competitive ELISA for Melamine Detection in Farm Products

To detect melamine(MEL) residue in foods and animal feeds,an indirect competitive ELISA(ic-ELISA) method was developed in this study based on preparation of monoclonal antibodies(MAbs) to MEL.The result showed that the best concentration of coating antigen is 1.0 μg/mL,the diluted MEL MAbs was 1∶1 000,the best blocking buffer was 50 g/L milk.The equation of linear regression is y =-15.12x + 107.3,coefficient correlation is R2=0.997.The intra-assay and inter-assay coefficients of variation were 1.07%-5.53% and 3.33%-11.92%,respectively.The method was utilized in the detection of MEL in pet foods,wheat flour tendon,animal feeds,milk and eggs with recovery ratio of 73.8%-84.32% for pet foods,67.3%-88.27% for wheat flour tendon,70.3%-85.9% for animal feeds,83.47%-89.76% for milk,and 76.2%-87.29% for eggs.Finally,The detection method developed by the optimal conditions,was highly specific and sensitive for detecting melamine.