血清载脂蛋白A5 ELISA法的建立及评价  

Development and evaluation of an ELISA for detecting serum apolipoprotein A5

作　　者：魏红霞[1] 张葵[1] 李雷[1] 倪军[1] 邱方[1] 顾光煜[1] 

机构地区：[1]南京大学医学院附属鼓楼医院检验科,南京210008

出　　处：《临床检验杂志》2009年第5期355-357,共3页Chinese Journal of Clinical Laboratory Science

基　　金：南京市科技发展重点项目(ZKM06055)

摘　　要：目的建立定量测定人血清中载脂蛋白A5(apoA5)浓度的ELISA双抗体夹心法,评价该法的敏感性、特异性、批内批间变异系数(CV),并分析apoA5与血脂指标的相关性。方法方阵法确定apoA5单克隆抗体的最佳检测浓度,建立apoA5标准曲线,ELISA双抗体夹心法测定重组apoA5蛋白,确定最低检测限及批内、批间CV,并检测300例正常人apoA5浓度和血脂指标,应用百分位数法计算apoA5的参考值。结果制备的apoA5单克隆抗体的最佳检测浓度为1∶800,最低检测限为2.12μg/L;特异性检测试验表明:apoA5单克隆抗体只针对重组apoA5蛋白和血清中天然的apoA5,apoA1和BSA的A值均<0.2;批内CV<3.5%,批间CV<5.5%。血清apoA5的参考值范围为255.92～383.81μg/L。相关性分析表明:apoA5与TG呈负相关(r=-0.27,P<0.05),与TC存在负相关趋势,与HDL-C呈正相关(r=0.36,P<0.05)。结论本实验室建立的ELISA双抗体夹心法灵敏度和特异性均较好,为apoA5的临床检测提供了可行的方法。Objective To establish an ELISA to quantitatively detect apolipoprotein A5 （ape A5 ） in serum and to analyze the correlation between apo A5 and serum lipid levels. Methods A double antibody sandwich ELISA for detecting ape A5 was developed based on the specific monoclonal antibody （mAb）. According to known concentrations of the recombinant ape AS, a reference curve was established. The low detection limit of the ELISA and the coefficient of variation （CV） values of the interassay and intra-assay were determined. Reference ape A5 ranges in healthy persons were analyzed by percentile method. Results The optimal concentration of ape 3_5 mAb for the ELISA was 1： 800. The low detection limit was 2.12 ng/ml. The specificity test showed that apoA5 mAb reacted only with the recombinant apo A5 and serum ape AS. The A values in measuring ape AI and BSA were 〈 0.2. The CV values of intra- and interassay were less than 3.5% and 5.5% respectively. The serum concentration of ape A5 in 300 healthy subjects was 255.92 - 383.81 μg/L. Dependability analysis showed that ape A5 was negatively correlated with TG （r = -0.27, P 〈 0.05 ） and TC, and positively correlated with HDL-C （ r = 0.36, P 〈 0.05 ）. Conclusions A highly sensitive and specific, ELISA for quantitation of ape A5 in serum has been established,which provides a practical way to measure ape A5.

关 键 词：栽脂蛋白A5 双抗体夹心酶联免疫吸附试验 

分 类 号：R446.1[医药卫生—诊断学]