降钙素原(PCT)单克隆抗体的制备及其在血清学检测中的应用

PREPARATION OF MONOCLONAL ANTIBODIES(MABS) OF PROCALCITONIN(PCT) AND ITS APPLICATION IN THE SEROLOGICAL DIAGNOSIS

以高纯度的PCT抗原免疫BALB/c小鼠,制备单克隆抗体并以此抗体作为包被抗体,通过包被液和封闭液的选择及条件优化,最终建立PCT双抗体夹心ELISA的血清学定量检测方法.建立的PCT双抗体夹心ELISA检测方法的灵敏度和特异性分别达到95.6%和97.8%,最佳的包被抗体浓度为400 ng/m L,酶标二抗的稀释度为1∶8 000,最适宜的缓冲液以及反应条件分别为包被液使用p H9.6的0.05 mol/L CB 4℃下包被12 h,封闭液使用0.6%BSA 37℃下封闭2 h.建立的PCT双抗体夹心法与罗氏试剂盒的定量检测结果呈现良好的相关性,是一种灵敏度高、特异性强、稳定可靠的血清学定量检测方法,为进行PCT的深入研究奠定了基础.

In order to establish a procalcitonin （PCT） double antibody sandwich ELISA quantitative detection method, high specific monoclonal antibodies （mAbs）, used as coating antibodies of PCT, were prepared after immunization of BALB/c mice with high purified PCT antigen and then selection and condition optimization of coating liquid and sealing liquid were accomplished, a serological quantitative detection method were developed. The sensitivity and specificity of double antibody sandwich ELISA quantitative detection method was 95.6% and 97.8% respectively. The most optimum coating antibody concentration was？400ng/mL, and the most favorable enzyme labeled secondary antibody dilution was 1：8000. The optimized buffer and reaction condition were coating buffer （PH=9.2 0.05 mol/L） at 4 ~C for 12h and sealing liquid （0.6% BSA） at 37 ℃ for 2 h respectively. The results detected by this method displayed considerable correlation with that of Roche quantitative detection method. The results indicated that double antibody sandwich ELISA quantitative method was a sensitive, specific and reliable detection method which provided basis for further study of PCT.