采用单克隆抗体夹心 ELISA 检测蓝舌病病毒群特异性抗原。
Detection of bluetongue virus group-specific antigen using monoclonal antibody based sandwich ELISA.
机构信息
Division of Virology, Indian Veterinary Research Institute, Mukteswar, 263138 Nainital Distt, Uttarakhand, India.
出版信息
Virol Sin. 2010 Dec;25(6):390-400. doi: 10.1007/s12250-010-3160-y. Epub 2010 Dec 21.
A monoclonal antibody (MAb) specific for the bluetongue virus (BTV) group specific antigen (VP7) was characterized for its reactivity with purified virus and recombinant BTV VP7 (rVP7) protein and its suitability for use in the sandwich ELISA. The MAb, designated as 5B5 was specific to VP7 and belongs to IgG2a subclass and was selected for the development of the sELISA in this study. The MAb had a titer of 1:25 with BTV and 1:2 with the rVP7 protein. The sELISA is based on capturing of BTV antigen with VP7 specific MAb followed by detection using BTV polyclonal antiserum raised in rabbits. The assay was evaluated with six cell culture adapted serotypes of BTV that have been isolated from India, 1, 2, 15, 17, 18 and 23. The assay could detect BTV antigen as early as day 8 in blood. It was also successfully applied for the detection of BTV group specific antigen in clinical samples of blood, washed RBCs, buffy coat and plasma. A total of 102 field samples from animals, suspected of being infected with BTV, were tested and 29.42% were positive. The blood samples were also amplified in cell culture which improved the sensitivity of the assay. Results confirmed that the sELISA is rapid and specific.
一种针对蓝舌病毒（BTV）群特异性抗原（VP7）的单克隆抗体（MAb）在与纯化病毒和重组 BTV VP7（rVP7）蛋白的反应性方面的特征，并评估其在夹心 ELISA 中的适用性。该 MAb 被指定为 5B5，特异性针对 VP7，属于 IgG2a 亚类，并被选为本研究中 sELISA 的开发。该 MAb 对 BTV 的效价为 1:25，对 rVP7 蛋白的效价为 1:2。sELISA 基于用 VP7 特异性 MAb 捕获 BTV 抗原，然后用兔多克隆抗血清检测。该测定方法用从印度分离的六种细胞培养适应的 BTV 血清型进行了评估，分别为 1、2、15、17、18 和 23。该测定方法可以在第 8 天检测到血液中的 BTV 抗原。它还成功地应用于临床血液、洗涤红细胞、白细胞层和血浆等临床样本中 BTV 群特异性抗原的检测。对来自疑似感染 BTV 的动物的 102 个现场样本进行了检测，其中 29.42%呈阳性。这些血液样本也在细胞培养中进行了扩增，提高了该测定方法的灵敏度。结果证实，sELISA 快速且具有特异性。
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