| Objective:Recent studies have found that PAPP-A was an inflammatory biomarker of coronary plaque instability. The level of serum PAPP-A in patients with acute coronary syndrome has some value in early diagnosis and prognosis. Maternal serum PAPP-A is different from non-pregnant serum PAPP-A present at low levels, however the methods of PAPP-A detection currently used with low sensitivity do not apply to detect non-pregnant serum. In our study, with oxygen channeling immunoassay, we established a quantitative analysis system for hypersensitivity level of serum PAPP-A in ACS patients, and evaluated whether this method was suitable for testing. On the basis, we test the specimens of ACS patients in different subgroups, furthermore laboratory studies whether PAPP-A in ACS patients is a good predictor of acute coronary syndrome or has the value of risk stratification and prognosis.Methods:Preparation of biotinylated PAPP-A receptor antibodies and antibody-coated microspheres combinated with serum PAPP-A by double-antibody-sandwich assay detect concentrations of PAPP-A by chemiluminescence analysis. Varying the loading mode and sample order, adjusting the ratio of the reagents and reaction time, temperature, etc., we optimized the reaction conditions of this detection method. We used ELISA Cale software to build a four-parameter calibration curve fitting equation, initially established homogeneous chemiluminescent immunoassay method to detect PAPP-A in hypersensitivity. Meanwhile we detected sensitivity, precision, accuracy, stability and other performance indicators of homogeneous chemiluminescent analysis method for PAPP-A, and reckoned the normal reference range. According to progression in patients with acute coronary syndrome we study to compare what was the difference PAPP-A levels in ACS patients and controls, as well as analysis other related indicators of ACS, discussed clinical correlation between the ACS and PAPP-A.Results:The detecting steps of PAPP-A: Add 25 ul biotinylated antibody(1: 2000), 25 ul acceptor beads(1: 100), 25 ul samples in 96 well plates, at 37℃ for 30 min. Add 175 ul donor beads in the dark conditions, at 37℃ for 10 min. Finally measure the signal value. The sensitivity of the assay we developed was 0.55 m U/L and its recovery rates were between 95%and 100%. The linearity, stability and specificity of the assay were convenient. The repeatability showed that the coefficient of variations(CVs) were not exceed 10% in the group, and the CVs were not exceed 15% between groups. The testing results were not subjected to interference by hemoglobin, triglycerides, or bilirubin. The assay was done the analysis associatively with TRFIA(r2=0.988,P<0.05). PAPP-A of 104 healthy serum samples(M/F: 43/61, Age: 24-69) were detected to establish the reference value, deducing the range were 1.60-25.12 m U/L.Conclusion:The assay we established to determine PAPP-A is sensitive and accurate, and the stability is excellent. There is greatly correlation between the assays we developed and the TRFIA. We furnish a basis for establishing the reference value of PAPP-A. The results demonstrate that the detection of PAPP-A meet the demand for clinical application. We analyze differences of ACS factors among groups, discuss and compare inflammatory markers related to ACS and their relevance. We preliminary hold that the detection of PAPP-A in hypersensitivity has value in risk stratification to ACS patients, but its mechanism of action and on ACS stratification and prediction of the standard requires further research and exploration. |
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