To establish a high chemiluminescence immune analysis method for the determination of C-reactive protein (CRP) in human serum.6-[N-(4-Aminobutyl)-N-ethylamino]-2,3-dihydro-1,4-phthalazinedione (ABEI) was used as luminous marker, magnetic beads was used as solid-phase reaction carrier. When AFP in serum combined with anti-CRP monoclonal antibody labeled with ABEI and another anti-CRP monoclonal antibody labeled with fluorescein isothiocyanate isomer I (FITC) to form the "sandwich" immune complex, then added magnetic beads coupled with anti-FITC monoclonal antibody. After washing procedure, adding the substrate, the chemiluminescence reaction of immune response was triggered, and the optical signal was generated. The relative intensity (RLU) of the reaction was measured by the instrument, and the concentration of the CRP in the sample was calculated. A complete test can be completed within 25 minutes, the average recovery rate was between 90% and 110%.A good linear relationship was detected in the concentration range of human serum CRP, the linear correlation coefficient of linear dilution effect is more than 0.9900. The method was accurate and reproducible, and the sensitivity of CRP was less than 0.13 ng/ml, and the detection limit was 0.13 ng/ml. The average recovery rate was 99%, the intra coefficient of variation (CV) was 3.24%~4.21%, and inter-assay coefficient of variation was 8.73%~9.35%. We established a chemiluminescence immunoassay to quantify the level of CRP in human serum. The results of performance evaluation showed it was an accurate, reliable and meet the needs of clinical testing.
Financed by the National Centre for Research and Development under grant No. SP/I/1/77065/10 by the strategic scientific research and experimental development program:
SYNAT - “Interdisciplinary System for Interactive Scientific and Scientific-Technical Information”.