摘要
目的:获得一株特异性强,灵敏度高的抗孕酮单克隆抗体,用辣根过氧化物酶标记孕酮-11α-羟基孕酮半琥珀酸酯制成酶标记物,建立酶联免疫分析(ELISA)方法,用于检测人血清中孕酮含量。方法:用免疫原11α-OH-孕酮-BSA免疫BALB/c小鼠,采用杂交瘤技术,得到14株抗孕酮单克隆抗体;用反射免疫分析(RIA)方法测定抗孕酮单克隆抗体的亲和常数和交叉反应率。结果:获得的14株抗孕酮单克隆抗体具有较高的亲和性和较小的价差反应率;建立ELISA方法学的灵敏度为0.12 ng/ml;批内变异系数为7.5%～11.4%;批间变异系数为:8.2%～12.1%;回收率93.8%～124.7%;高值临床样品系列倍比稀释后,测定值与稀释度呈线性相关,相关系数为r=0.995 7;与全自动化学发光测定的临床样品值比对结果较好,线性方程为y=0.789 4x+0.760 0,r=0.912 6。结论:方法学鉴定结果符合免疫分析的基本要求。
Objective：The study was aimed to get highly specific and sensitive against progesterone monoclonal antibodies. The enzyme conjugate was prepared by labeling progesterone-llα-hemisuccinate （P-1lα-His） with Horseradish Peroxidase （HRP） to form P-1lα-His-HRP, development a highly specific and sensitive Enzyme Linked Immunosorbent Assay （EL1SA） to measm＇e progesterone in human sellun. Methods： The BALB/c mice were irmnunized using the antigen that was the conjunction 1lα-OH-Progesterone-BSA by hybridoma technique. The Affinity Constant and Rate of Cross Reactions were determined by Radioirrununoassay （RIA）. Results： 14 strains against progesterone monoclonal anti- bodies were acquired. The ELISA to measure progesterone in human serum was developed. The standard range of the mefllod was 0.3-100 ng/ml. The assay sensitivity was 0.12 ng/ml. The intra-assay and inter-assay coefficients of variance （CVs） varied was 7.5 %-11.4 % and 8.2 %- 12.1%. Analytical recover）, was 93.8%-124.7%. Compared with detected value clinically, the correlative equation was y = 0. 789 4x ＋ 0. 760 0, r = 0. 912 6. Conclusion： Methodology appraisal results with the basic requirements of immune analysis.
出处
《中国免疫学杂志》
CAS
CSCD
北大核心
2011年第6期548-551,555,共5页
Chinese Journal of Immunology
