Abstract: AIM: To establish a sandwich ELISA for quantitative measurement of supersensitivity C-reactive 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.

Key words: quantitative measurement, hs-CRP, sandwich ELISA