甲基睾酮的控温相分离免疫分析

Phase Separation Immunoassay of Methyltestosterone

建立了一种新的甲基睾酮（MT）测定的免疫分析方法。通过将MT与牛血清白蛋白（BSA）偶联制得了MT全抗原MT-3－BSA（AgMT）。将AgMT经兔子免疫获得了针对MT的多克隆抗体（AbMT）。与温度敏感性水凝胶偶联的抗原（PNIP-AgMT）和游离半抗原MT竞争性地与有限量的AbMT和辣根过氧化物酶（HRP）的偶联物（AbMT－HRP）反应。根据临界析相温度以上免疫复合物沉淀的特性进行分离，根据沉淀相HRP催化H_2O_2氧化4-氨基安替比林显色反应的初速度可确定样品中MT的浓度。该法对MT测定的线性范围在50μg/L^7．5mg/L最低检出限为50μg/L。MT在人血清中的回收率比较满意。

A notel immunoassay of methyltestosterone (MT) in serum was developed. The antigen of MT (AgMT) was synthesized by covalently bonding MT to bovine serum albumin (BSA). The polycolnal antibody of AgMT (AbMT ) was raised from rabit. AgMT was conjugated to poly-N-isopropylacrylamide (PNIP) to form AgMT bound thermally reversible hydrogel PNIP-AgMT. A competitive immunoassay method based on the competition of PNIP-AgMT and free MT with limited amount of horseradish peroxidase(HRP) labeled AbMT was established. The separation of PNIP in the assay process was achieved by precipitation of the immuno-complex above its critical solution temperature. The amount of MT was determined by measuring the initial rate of the oxidation of 4-aminoantipyrine by H2O2 catalyzed by HRP-immuno- complex precipitated. The linear range for MT was 50 μg/L ~ 7. 5 mg/L and the detection limit was 50 μg/L. The recoveries of MT form human serum were satisfactory.