Saliki J T, Libeau G, House J A, Mebus C A, Dubovi E J
Foreign Animal Disease Diagnostic Laboratory, U.S. Department of Agriculture, Greenport, New York 11944.
J Clin Microbiol. 1993 May;31(5):1075-82. doi: 10.1128/jcm.31.5.1075-1082.1993.
A blocking enzyme-linked immunosorbent assay (B-ELISA), using two neutralizing monoclonal antibodies (MAbs), was established and compared with the virus neutralization test (VNT) for detecting specific peste-des-petits-ruminants virus (PPRV) antibody in caprine and ovine sera. This technique was developed because VNT, the only available specific serological test for PPRV and the cross-reactive rinderpest virus (RPV), is time-consuming and unaffordable for most laboratories in regions where both peste des petits ruminants and rinderpest occur. The test depends on the blocking of the binding of the MAb to a specific epitope in the presence of positive serum. Test conditions were optimized by using peste-des-petits-ruminants and rinderpest sera that were known to be VNT positive and negative. A blocking format, in which serum is preincubated with a solid-phase PPRV antigen and then incubated with the MAb, yielded levels of sensitivity and specificity superior to those of a competitive format, in which the two reagents are added simultaneously. A threshold value of 45% inhibition, representing the mean for a negative population (n = 277) plus 2.7 standard deviations, was adopted for routine screening. A total of 605 serum samples were screened by B-ELISA and the VNT. The sensitivity and specificity of B-ELISA relative to the VNT were 90.4 and 98.9%, respectively. Of 264 field serum samples tested, 11 (4.2%) could not be assayed by the VNT because of contamination or cytotoxicity; the overall agreement quotient between results of the two tests (n = 253) was 0.91. A high correlation (r>/=0.98) was observed between B-ELISA and the VNT for endpoint titration of sera (n=57). Because B-ELISA proved to be nearlyas sensitive and specific as the VNT while being simpler and more rapid, it would be an adequate substitute for the VNT for assessing herd immune status and for epidemiologic surveillance.
建立了一种使用两种中和单克隆抗体（MAb）的阻断酶联免疫吸附测定（B-ELISA），并将其与病毒中和试验（VNT）进行比较，以检测山羊和绵羊血清中特异性小反刍兽疫病毒（PPRV）抗体。开发这项技术是因为VNT是唯一可用于检测PPRV和交叉反应性牛瘟病毒（RPV）的特异性血清学检测方法，但对于小反刍兽疫和牛瘟都流行的地区的大多数实验室来说，它既耗时又成本高昂。该检测依赖于在阳性血清存在下阻断MAb与特定表位的结合。通过使用已知VNT阳性和阴性的小反刍兽疫和牛瘟血清优化了检测条件。一种阻断形式，即血清先与固相PPRV抗原预孵育，然后与MAb孵育，其灵敏度和特异性水平优于两种试剂同时加入的竞争形式。采用45%抑制阈值作为常规筛查标准，该阈值代表阴性群体（n = 277）的平均值加上2.7个标准差。通过B-ELISA和VNT共筛查了605份血清样本。B-ELISA相对于VNT的灵敏度和特异性分别为90.4%和98.9%。在检测的264份现场血清样本中，有11份（4.2%）由于污染或细胞毒性无法通过VNT检测；两种检测结果（n = 253）之间的总体一致性商数为0.91。在血清终点滴定（n = 57）中，观察到B-ELISA与VNT之间具有高度相关性（r≥= 0.98）。由于B-ELISA被证明与VNT几乎一样灵敏和特异，同时更简单、更快速，它将是评估畜群免疫状态和进行流行病学监测时VNT的合适替代方法。
