钙调素ELISA定量检测方法的建立与实验条件优化  

Development and optimization of quantitative ELISA for calmodulin

作　　者：王娟[1] 李晓军[1] 潘继文[2] 虞伟[1] 廖建辉[3] 赵慧斌[1] 王艾丽[1] 武建国[1] 

机构地区：[1]南京军区南京总医院解放军医学检验中心临床免疫实验室,江苏南京210002 [2]东南大学国家教委分子与生物分子电子学开放实验室,江苏南京210096 [3]合肥市第一人民医院检验科,安徽合肥230061

出　　处：《医学研究生学报》2003年第6期401-403,406,F002,共5页Journal of Medical Postgraduates

基　　金：江苏省"13 5工程"重点人才基金资助课题

摘　　要：目的 :　研究ELISA法定量测定钙调素 (CaM)的最优条件。　方法 :用重组人CaM、兔抗CaM ,通过对聚苯乙烯反应板进行紫外辐照处理 ,改善抗原固相化条件 ;通过对抗原包被液、包被时间等条件优化 ,建立简便、快速、准确的ELISA定量技术。　结果 :以pH 7.4 0 .0 1mol/L的PB为包被液 ,包被及后续封闭时间为 4℃ ,静置 72h ,CaM包被浓度为 5 μg/ml,抗原抗体反应时间为 37℃ 6 0min ,可获最佳CaM定量结果。　 结论 :建立检测CaM的ELISA测定方法 ,敏感性、重复性均在临床检测可接受的范围内 ,标准曲线的线性也较好 。Objectives:To optimize the procedures of quantitative determination of calmodulin(CaM) by ELISA. Methods:By using recombinant human CaM as coating antigen, the polystyrene inicrotiter plates was treated with ultraviolet radiation to improve the immobilization of short peptides. The constitution of buffer and the coating time were optimized as well. Results:The optimized experimental conditions were as follows: The coating solution was 0.01 mol/L, pH 7.4 PB. The coating and blocking condition was 4℃, 72 h. The coating concentration of CaM was 5 μg/ml and the antigen antibody reaction condition was 37℃, 60 min. The standard curve of the quantitative ELISA for CaM assay was satisfactory. The sensitivity and repetition of the method were all in the acceptable range of clinical analysis. Conclusions:A simple, rapid and precise ELISA method for CaM quantitative analysis was developed. It could be used in quantitative study of inner or extra cellular CaM.

关 键 词：钙调素 ELISA法 检测 

分 类 号：R392.33[医药卫生—免疫学]