Objective: To develop a kit of time-resolved fluoroimmunoassay (TRFIA) for detection of Schistosoma japonicum protein SjP38, and evaluate its effectiveness.
Methods: The anti 9G7 SjP38 monoclonal antibody was used as the capture anti-body coated with 96-hole plate, and the Eu3+ labeled 1A6 monoclonal antibody was used as the detection antibody to establish the TRFIA SjP38 kit. In addition, the accuracy, sensitivity, precision, stability and coincidence rate to pathogenic diagnosis of the kit were evaluated.
Results: This established kit possessed high accuracy, wide linear range from 2 to 1 250 ng/ml, high sensitivity with the minimum detectable concentration of 0.14 ng/ml, and good precision (the coefficient variation of the intra- and inter-assay were 3.6% to 4.6% and 5.1% to 6.7%, respectively). The stability tests showed that the reagents could be stable for six months at 4 ℃, 7 d at 37 ℃. The positive and negative corresponding rates to the pathogen detection method were 95% and 100% respectively.
Conclusions: All the performance and detection indicators of the kit have reached the requirements of clinical test, but its clinical application still needs further validation.
［摘要］ 目的 研制日本血吸虫SjP38的时间分辨免疫荧光分析 (Time-resolved fluoroimmunoassay, TRFIA) 试剂盒, 并评 价其效能。 方法 将抗SjP38的9G7单克隆抗体作为捕获抗体包被96孔板, 用Eu3+标记1A6单克隆抗体作为检测抗体, 研制SjP38 TRFIA试剂盒, 并评价试剂盒的准确度、灵敏度、精密度、稳定性以及与病原学检测方法的符合率等指标。结 果自制试剂盒准确度好, 线性范围为2～1 250 ng/ml, 最低检出浓度为0.14 ng/ml; 精密度良好, 分析内精密度为3.6%～4.6%, 分析间精密度为5.1%～6.7%。定性试验表明该试剂盒可在4 ℃稳定半年, 37 ℃稳定7 d。自制试剂盒与病原学检 测方法的阳性符合率为95%, 阴性符合率为100%。 结论 所研制试剂盒的各项性能及检测指标均达到临床检验的要 求, 但临床推广使用还有待进一步的验证。.
Keywords: Double-antibody sandwich; Recombinant protein SjP38; Schistosoma japonicum; Time-resolved fluoroimmunoassay (TRFIA).