Development Of Luminescent Oxygen Channeling Immunoassay For PAPP-A

Objective:Based on luminescent oxygen channel immunoassay(LOCI) of AlphaScreen platform, we develope a homogeneous luminescence immunoassay system for pregnancy associated plasm a protein A(PAPP-A) detection and make a evaluation of methodology and clinical. Methods:Rabbit polyclonal antibody for PAPP-A and mouse monoclonal antibody for PAPP-A coated receptor microsbeads, biotm, respectively; composed of two antibodies,titration is used to determine the best combination;Titration method is used to determine the optimal concentration of the receptor microbeads and antibody; biotin and buffer of the detection system,reagent and sample order, and the proportion of reagent and sample and two phase reaction time were optimized. Based on optimizing the four parameter fitting equation of standard curve, according to the standard curve can be unknown concentration in clinical specimens by emitting signal value of PAPP-A concentration. To detect the clinical specimens according to the curve and make methodology and clinical evaluation. Study by ELISA Calc software for statistical analysis. Results:The monoclonal antibody of PAPP-A was labeled with biotin,the polyclonal antibody of PAPP-A was coated on receptor particles.The LOCI reagents also contained sensitizer particles coated with streptavidin.The optimal test conditions and analytical performance of the method were studied. The within-run and the between-run coefficients of variation were 5.91%~7.94%and 6.14%~9.69%,respectively; the analytical sensitivity was 2.8mU/L and the function sensitivity was 4.6mU/L,good linear in 2.8mU/L-8000mU/L range.;the recovery rate was 96.7%-100.3%. The interference rate of hemolysis,icterus and triglycerides were less than 10%; there is no Hook effect of PAPP-A concentrations up to 8000mU/L;the correlation coefficient between clinical samples detection results and Time resolved fluoroisnmuno assay analysis results was 0.989. Conclusion:This system can be used for quantitative detection of serum PAPP-A,and with no washing, strong sensitivity, high accuracy, good repeatability, easy operation etc,and further research is expected to promote the clinical application.