Asia1型口蹄疫病毒胶体金免疫层析检测方法的建立

Establishment of colloidal gold-immunochroma-togrphy assay strip for detection of type Asia1 foot-and-mouth disease virus

摘要:

为建立一种快速、简便、灵敏检测Asia1型口蹄疫病毒的胶体金免疫层析方法(GICA)。本研究采用柠檬酸三钠还原法制备胶体金颗粒, 标记纯化的抗Asia1型口蹄疫病毒的单克隆抗体, 将该标记物与羊抗豚鼠IgG分别包被在硝酸纤维素膜(Nitrocellulose membrane)上, 作为检测带和质控带。经条件优化, 组装成检测Asia1型口蹄疫的诊断试纸条。用该试纸条分别对A、O、C和Asia1型口蹄疫病毒抗原以及猪水泡病病毒抗原等87份样品进行了检测, 发现该试纸条不与口蹄疫病毒A、O、C型以及猪水泡病病毒抗原发生反应, 特异性良好。用该试纸条对口蹄疫细胞毒(TCID50为6.25)的10倍系列稀释液进行了检测, 最低可以检测到大约10-4。该试纸条与其他传统诊断方法的符合率为98.8%。初步实验确定该试纸条在4oC下可保存3个月、37oC和室温下大概可保存1周左右。该试纸条是一种快速、灵敏、特异的FMD抗原检测方法, 对现场检测具有一定实用价值。

Abstract:

To establish a sensitive, rapid and simple gold immunochromatography assay (GICA) for detecting Asia1 type of foot-and-mouth disease virus (FMDV) from the field samples. The purified anti-FMDV type Asia1 monoclonal antibody labeled with collidal gold and the goat anti-Guinea pig IgG were wrapped onto nitrocellulose membrane as the test line (T line) and the control line (C line), respectively. The strip was then further optimized. A total of 87 field samples were detected. The results indicated a correct rate of 98.8% for detecting FMDV Asia1 type. No cross reaction was found with swine vesicular disease (SVD) and FMDV O, A and C type antigen. The sensitivity of the strip can reach to 10-4 (TCID50 6.25). It had the same results for positive and negative specimens tested in three times. This strip could be stored at 4oC for three months. In this study, the established gold immunochromatographic strip test kit is simple, rapid, sensitive and specific for detecting FMDV type Asia1, and is potentially useful for the for pen-side diagnosis.