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

Detection of Yersinia pestis-specific F1 antigen by a double monoclonal antibody sandwich enzyme-linked immunosorbent assay

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（x。=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），～致性为96．99％[1／4×（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＋273／2844-246／257＋246／249）]，Youden指数为0．9492。DMcAbS—ELISA法对鼠疫菌检测灵敏度为2．7×10^4cfu／ml，RIHA法为2．2×10^5cfu／ml；两种方法检测F1抗原灵敏度均为10μg／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, DMeAbS-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 germieulture, DMeAbS-ELISA and RIHA, and the difference was not statistically significant（x2 = 5.65, P 〉 0.05）. The coincidence rate of DMeAbS-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 x （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 x （273/276 ＋ 273/284 ＋ 246/257 ＋ 246/249）J and 0.9492, respectively, for RIHA. The detection sensitivity of DMcAbS-ELISA and RIHA was 2.7 ×10^4 cfu/ml and 2.2 ×10^5 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.