ELISA检测方法中最佳封闭液和样品稀释液的筛选研究

摘要

[目的]为提高酶联免疫吸附法(ELISA)的敏感性和特异性,研究分析了不同类别和不同浓度的封闭液和样品稀释液对ELISA检测结果的影响.[方法]研究采用酪蛋白(Casein)、明胶(Gelatin)、BSA、山羊血清(GS)、马血清(HS)和兔血清(RS)等不同类封闭液和同一类不同浓度封闭液进行ELISA检测试验.[结果]2％BSA较1％BSA和3％BSA封闭效果好,2％ Casein和1％ Casein较3％Casein封闭效果好,8％RS和10％RS封闭效果强于6％RS和7％RS；与BSA和Casein相比,RS具有更好的封闭效果,且8％RS封闭液和8％RS样品稀释液组合最佳.[结论]良好的封闭液和样品稀释液组合可有效降低非特异性反应,提高ELISA检测方法的敏感性和特异性,这为良好ELISA检测方法的建立提供了重要实践指导作用.%[Objective] This study aimed to increase the sensitivity and specificity of enzyme-linked immunosorbent assay (ELISA) through analyzing the effects of different blocking buffers and sample diluents and their different concentrations on the result of ELISA.[Method] Different types of blocking buffer [casein,gelatin,BSA,goat serum (GS),horse serum (HS) and rabbit serum (RS)]and sample diluent (PBST,casein,gelatin,BSA,GS,HS and RS) as well as their different concentrations were tested in ELISA to screen the optimal combination of blocking buffer and sample diluent.[Result] The results showed that 2％ BSA had better effect on blocking than 1％ and 3％ BSA,and both 2％ and 1％ casein had better blocking effect than 3％ casein； 8％ and 10％ RS showed better blocking effects than 6％RS and 7％RS； compared to BSA and casein,· RS had the best effect on blocking,and 8％RS performed best as the blocking buffer and sample diluent.[Conclusion] A good combination of blocking buffer and diluent can effectively reduce the non-specific reaction and improve the sensitivity and specificity of ELISA.This study provides an important reference for the development of a perfect ELISA method.