基于Tris辅助法制备猪轮状病毒胶体金免疫层析试纸条

Preparation of Porcine Rotavirus Colloidal Gold Immunochromatographic Strips Based on Tris Assisted Method

基于Tris辅助法制备颗粒大小均匀、颜色鲜艳、稳定性好的胶体金,并以此建立一种快速、便捷、稳定的猪轮状病毒现场检测技术。试验采用Tris辅助法制备胶体金,将其与轮状病毒标记抗体Ab2偶联,喷涂在结合垫上,以轮状病毒捕获Ab1抗体和羊抗鼠IgG包被硝酸纤维素膜,分别作为检测线和质控线。同时优化胶体金最适pH、抗体最佳使用量,并通过灵敏度试验、特异性试验、重复性试验、稳定性试验以及临床试验,考察试纸条的性能。结果表明,基于Tris辅助法制备的胶体金猪轮状病毒免疫层析试纸条,可检出稀释400倍临床样品中的轮状病毒,与相似病毒无交叉干扰,批间、批内试纸条检测同一样品时,无明显差异,在4℃、常温和37℃保存40 d后检测结果稳定。说明基于Tris辅助法制备胶体金的猪轮状病毒免疫层析试纸条,各方面性能均达到临床快速检测的要求,可用于临床检测。

Based on the Tris-assisted method,the colloidal gold with uniform particle size,bright color and good stability was prepared,and a rapid,convenient and stable immunochromatographic technology was established for the on-site detection of porcine rotavirus based on the Tris-colloidal gold.The colloidal gold was coupled with rotavirus labeled antibody(Ab2)and sprayed on conjugating pad.The rotavirus capture antibody(Ab1)and goat anti-mouse IgG were coated on nitrocellulose(NC)membrane as the detection line and the quality control line,respectively.At the same time,the optimal pH of colloidal gold solution and the optimal amount of antibody were optimized.The sensitivity,specificity,stability,repeatability and validity of the test strips was analyzed.The results showed that the colloidal gold immunochromatographic test strip can detect rotavirus in 400 times diluted clinical samples,without cross reacting with similar viruses.No obvious difference was observed between different strips from the inter batch and intra batch test results,and it can be stored at 4℃,normal temperature and 37℃for 40 days.It shows that the colloidal gold immunochromatographic test strip of porcine rotavirus based on the Tris assisted method meets the requirements of rapid clinical detection in all aspects.It is suitable for clinical detection.