Raj G Dhinakar, Rajanathan T M C, Kumar C Senthil, Ramathilagam G, Hiremath Geetha, Shaila M S
Department of Animal Biotechnology, Madras Veterinary College, Tamil Nadu Veterinary and Animal Sciences University, Chennai, India.
Vet Microbiol. 2008 Jun 22;129(3-4):246-51. doi: 10.1016/j.vetmic.2007.11.026. Epub 2007 Dec 3.
Peste des petits ruminants (PPR) is one of the most economically important diseases affecting sheep and goats in India. An immunofiltration-based test has been developed using either mono-specific serum/monoclonal antibodies (mAb) prepared against a recombinant truncated nucleocapsid protein of rinderpest virus (RPV) cross-reactive with PPR virus. This method consists of coating ocular swab eluate from suspected animals onto a nitrocellulose membrane housed in a plastic module, which is allowed to react with suitable dilutions of a mAb or a mono-specific polyclonal antibody. The antigen-antibody complex formed on the membrane is then detected by protein A-colloidal gold conjugate, which forms a pink colour. In the immunofiltration test, concordant results were obtained using either PPRV mAb or mono-specific serum. Another test, an antigen-competition ELISA which relies on the competition between plate-coated recombinant truncated 'N' protein of RPV and the PPRV 'N' protein present in ocular swab eluates (sample) for binding to the mono-specific antibody against N protein of RPV (in liquid phase) was developed. The cut-off value for this test was established using reverse transcription polymerase chain reaction (RT-PCR) positive and negative oculo-nasal swab samples. Linear correlation between percent inhibition (PI) values in antigen-competition ELISA and virus infectivity titres was 0.992. Comparison of the immunofiltration test with the antigen-competition ELISA yielded a sensitivity of 80% and specificity of 100%. These two tests can serve as a screening (immunofiltration) and confirmatory (antigen-competition ELISA) test, respectively, in the diagnosis of PPR in sheep or goats.
小反刍兽疫（PPR）是印度影响绵羊和山羊的最重要的经济疾病之一。已开发出一种基于免疫过滤的检测方法，该方法使用针对与PPR病毒交叉反应的牛瘟病毒（RPV）重组截短核衣壳蛋白制备的单特异性血清/单克隆抗体（mAb）。该方法包括将疑似动物的眼拭子洗脱液包被在塑料模块中的硝酸纤维素膜上，使其与合适稀释度的mAb或单特异性多克隆抗体反应。然后通过蛋白A-胶体金缀合物检测膜上形成的抗原-抗体复合物，其会形成粉红色。在免疫过滤试验中，使用PPRV mAb或单特异性血清可获得一致的结果。还开发了另一种检测方法，即抗原竞争ELISA，该方法依赖于板包被的RPV重组截短“N”蛋白与眼拭子洗脱液（样品）中存在的PPRV“N”蛋白竞争结合液相中针对RPV N蛋白的单特异性抗体。使用逆转录聚合酶链反应（RT-PCR）阳性和阴性眼鼻拭子样本确定了该检测的临界值。抗原竞争ELISA中的抑制百分比（PI）值与病毒感染性滴度之间的线性相关性为0.992。免疫过滤试验与抗原竞争ELISA的比较得出敏感性为80%，特异性为100%。这两种检测方法可分别作为绵羊或山羊PPR诊断中的筛查（免疫过滤）和确证（抗原竞争ELISA）检测方法。
