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目的:制备兔抗去氢甲睾酮(DMT)抗体,为采用免疫法分析食品中该激素的残留提供基础。方法:采用碳二亚胺法将肟化后的DMT分别偶联到钥孔血蓝蛋白、鸡卵清蛋白和牛血清白蛋白蛋白载体上,制备完全抗原,并利用薄层层析法、红外和紫外光谱法对其进行确证后,免疫新西兰大白兔;通过间接ELISA法测定和比较2种兔抗血清的效价、IC50及交叉反应率。结果:成功制备了DMT的完全抗原,其抗体效价分别为1∶64000(KLH-DMT)和1∶256000(OVA-DMT),IC50值分别为0.24μg/L和5.29μg/L;抗血清与丙酸睾酮酯、群勃龙的IC50值均大于781.25μg/L。结论:制备的DMT完全抗原具有较高的免疫原性,抗体与类似物丙酸睾酮酯、群勃龙无明显的交叉反应,可用于ELISA实验。
OBJECTIVE: To prepare rabbit anti-dehydrogenated methyltestosterone (DMT) antibody and provide the basis for the immunoassay to analyze the residue of this hormone in food. Methods: DMTs were coupled to keyhole limpet hemocyanin, chicken ovalbumin and bovine serum albumin protein carrier respectively by carbodiimide method to prepare the complete antigen. Thin layer chromatography, infrared and New Zealand white rabbits were immunized by UV spectrophotometry. The titer, IC50 and cross-reactivity of the two rabbit antisera were measured and compared by indirect ELISA. Results: The complete antigen of DMT was successfully prepared with antibody titers of 1:64000 (KLH-DMT) and 1:256000 (OVA-DMT) respectively, and the IC50 values were 0.24μg / L and 5.29μg / L, respectively. And testosterone propionate, the value of TBBT IC50 values were greater than 781.25μg / L. CONCLUSION: The prepared DMT antigen has high immunogenicity. The antibody has no obvious cross-reaction with testosterone propionate and trabupram and can be used in ELISA.