双单克隆抗体夹心酶联免疫吸附试验检测鼠疫F1抗原的实验观察

摘要

目的 观察双单克隆抗体(F1-McAb)夹心酶联免疫试验(DMcAbS-ELISA)快速检测鼠疫F1抗原的敏感性和特异性.方法 采用鼠疫细菌学检验、DMcAbS-ELISA和反向间接血球凝集试验(RIHA)对比检测鼠疫感染鼠和阴性对照鼠脏器标本.结果 共检测225份阴性对照鼠脏器标本,鼠疫细菌学检验、DMcAbS-ELISA和RIHA法检测F1抗原均为阴性.共检测308只鼠疫感染鼠脏器标本,鼠疫细菌学检验、DMcAbS-ELISA、RIHA法阳性率分别为92.21％(284/308)、90.91％(280/308)和89.61％(276/308),3种方法比较,差异无统计学意义(x2=5.65,P＞0.05).DMcAbS-ELISA法与鼠疫细菌学检验结果符合率为97.00％[(274+243)/533],Kappa值为0.940;与RIHA法符合率为99.25％[(276+253)/533],Kappa值为0.985.脏器标本F1抗原检测的真实性比较:DMcAbS-ELISA法敏感性为96.48％(274/284),特异性为97.59％(243/249),阳性预测值为97.86％(274/280),阴性预测值为96.05％(243/253),一致性为96.99％11/4×(274/280+274/284+ 243/253+243/249)|,Youden指数为0.9407；RIHA法的敏感性为96.13％(273/284),特异性为98.80％(246/249),阳性预测值为98.91％(273/276),阴性预测值为95.72％(246/257),一致性为97.39％[1/4×(273/276+273/284+246/257+246/249)],Youden指数为0.9492.DMcAbS-ELISA法对鼠疫菌检测灵敏度为2.7×104cfu/ml,RIHA法为2.2×105 cfu/ml;两种方法检测F1抗原灵敏度均为10 μg/L.结论 DMcAbS-ELISA法检测鼠疫F1抗原具有敏感、特异、简便、快速的特点,是有应用价值的鼠疫快速诊断技术.%Objective To study the sensitivity and specificity of a double monoclonal antibody sandwich enzyme-linked immunosorbent assay (DMcAbS-ELISA)for the detection of F1 antigen of Yersinia pestis (Y.pestis).Methods Viscera (viz.liver and spleen)specimens of infected mice with virulent Y.pestis and negative control mice were detected by bacteriological test,DMcAbS-ELISA and reverse indirect hemagglutination assay (RIHA) for the F1 antigen.Results The 225 control specimens were all negative tested by plague bacteriology testing,DMcAbS-ELISA and RIHA.A total of 308 plague-infected mouse organ specimens were tested,and the positive detection rate was 92.21％ (284/308),90.91％(280/308) and 89.61％ (276/308),respectively,with germiculture,DMcAbS-ELISA and RIHA,and the difference was not statistically significant(x2=5.65,P＞0.05).The coincidence rate of DMcAbS-ELISA and bacterial culture was 97.00％[(274+243)/533],Kappa =0.940;RIHA in line with the rate was 99.25％[(276+253)/533],Kappa =0.985.Authenticity comparison of F1 antigen detection in viscera specimens:sensitivity,specificity,positive predictive value,negative predictive value,adjusted agreement and Youden's index was 96.48％(274/284),97.59％(243/249),97.86％ (274/280),96.05 ％(243/253),96.99％[1/4×(274/280+274/284+243/253+243/249)]and 0.9407,respectively,for DMcAbS-ELISA and 96.13％(273/284),98.80％(246/249),98.91％(273/276),95.72％(246/257),97.39％[1/4×(273/276+273/284+246/257±246/249)]and 0.9492,respectively,for RIHA.The detection sensitivity of DMcAbS-ELISA and RIHA was 2.7×104 cfu/ml and 2.2×105 cfu/ml,for Y.pestis,respectively,and was 10 μg/L for F1 antigen.Conclusions DMcAbS-ELISA assay is a sensitive,specific,simple and fast method for detection of the F1 antigen,and it has a potential application value in rapid diagnosis of plague.