摘要
目的建立小鼠肝炎病毒（MHV）双抗体夹心ELIsA检测方法。方法将4株特异性抗MHVN蛋白的单克隆抗体进行抗体配对试验确定3D4H9为捕获抗体，以辣根过氧化物酶（HRP）标记的4G9F2为检测抗体；通过方阵试验确定了3D4H9和4G9F2-HRP的最佳工作浓度分别为12．8ng／mL和70．84ng／mL，以P／N〉2，同时D450nm≥0．248作为阳性临界值的判定。结果该ELISA方法重复性变异系数小于10％，与小鼠细小病毒、仙台病毒、小鼠肺炎病毒、呼肠孤病毒3型等无交叉反应，对358份血清样品进行检测，与美国EBI公司的间接ELISA试剂盒检测结果无明显差异。结论本研究建立的双抗体夹心ELISA方法检测MHV具有较高的特异性和敏感性，可以用于MHV的病原学检测。
Objective To develop a sandwich ELISA for detection of mouse hepatitis virus （MHV）. Method The ELISA was standardized by pair-matching experiment, and the working concentrations of the mAb 3D4H9 and HRP-conjugate mAb 4G9F2 were 12.8 ng/mL and 70.84 ng/mL, respectively, and judging with P/N〉2 and absorbance450nm ≥ 0.248 as positive criteria. Results The DAS-ELISA was specific for detection of MHV, but no cross-reaction with minute virus of mouse, reovirus type 3, sendai virus, pneumonia virus of mice. The intra-assay and inter-assay coefficient of variability were within 10%. In addition, a total 358 samples were tested by the DAS-ELISA, and indirect ELISA, there was no significant difference between results. Conclusion The DAS-ELISA was sensitive and specific which provided a useful tool for diagnosis of MHV.
出处
《实验动物与比较医学》
CAS
2015年第1期6-9,36,共5页
Laboratory Animal and Comparative Medicine
基金
上海市科委科技创新行动计划（08140901200）
