中国临床药理学与治疗学2009，Vol.14Issue(1)：84-89,6.
定量检测人超敏C-反应蛋白双抗体夹心ELISA方法的建立及初步临床应用
Establishment of a sandwich ELISA for quantitative measurement of human supersensitivity C-reactive protein and primary clinical applica-tion study
摘要
Abstract
AIM: To establish a sandwich ELISA for quantitative measurement of supersensitivity C-reac-tive protein(hs-CRP). METHODS: Anti-CRP monoclonal antibodies (mAbs) prepared by our laboratory were purified by Protein A affinity chromatography and analyzed by SDS-PAGE and Western-blotting to test their characteristics. All the mAbs were labeled with horseradish peroxidase by sodium oxidation method and antibody mating test were performed using anti-CRP mAb as coating antibody and HRP labeled anti-CRP mAb as labeled antibody, in which optimal concentrations were defined by square mateix titration. Standard curve was performed using purified CRP and the sensitivity, reproducibility and recovery rate test of ELISA was evaluated. The CRP levels in plasma were measured in this assay. RESULTS: The optimal paired antibodies were anti-CRP mAb 1C10 and HRP labeled anti-CRP mAb 2C11, and the optimal concentrations were 10 μg/mL and 1: 2000,respectively. The coefficient of variation were 3.1% to 9.7 % within assay and 3.6% to 13.6% between assay. The sensitivity in this assay was 8.3 ng/mL. The recovery rate was 90% to 109%. According to the result,68 normal persons' hs-CRP level in plasma of coronary arteriography were detected by ELISA, 59 with stenosis < 50% and 67 with stenosis ≥50%. The results showed that the plasma hs-CRP level in patients with stenosis < 50% (3.65 ± 1.15) mg/L was significantly higher ( P < 0.05)than hs-CRP in normol persons(1.75 ± 0.74) mg/L, the plasma hs-CRP level in stenosis ≥ 50% patients ( 8.93 ± 3.29) mg/L was significantly higher (P < 0.05) than stenosis < 50% patients. CONCLUSION: A sandwich ELISA for detecting hs-CRP was established.关键词
定量检测/超敏C-反应蛋白/ELISA双抗体夹心法Key words
quantitative measurement/hs-CRP/sandwich ELISA分类
医药卫生引用本文复制引用
沈丹丹,卞智萍,何国平,顾春荣,徐晋姚,杨笛,张寄南..定量检测人超敏C-反应蛋白双抗体夹心ELISA方法的建立及初步临床应用[J].中国临床药理学与治疗学,2009,14(1):84-89,6.基金项目
重大疾病分子诊断和生物治疗的高技术平台基金项目(XK200705) （XK200705）
