三聚氰胺免疫亲和柱的制备和测试

Preparation and Evaluation of Melamine Immunoaffinity Column

制备高容量的三聚氰胺免疫亲和层析柱,以识别三聚氰胺的单克隆抗体为配体,采用硅烷化方法,对抗体进行包埋,装填小柱。采用三聚氰胺系列标准溶液进行富集试验,通过紫外光谱(三聚氰胺最大紫外吸收波长240 nm)和液相色谱-质谱测试流出液和洗脱液中三聚氰胺的浓度,确定了免疫亲和柱的柱容量为16.47μg。试验中对亲和小柱的润洗溶液、上样溶液、洗脱溶液和再生溶液进行了优化,结果表明,该免疫亲和层析柱的富集效率可达66倍。以回收率高于60%计,该柱的再生次数至少为5次。

An immunoaffinity column （IAC）for melamine with high eolumn capacity was prepared in this study. Based on a monoclonal antibody against melamine,a silanizati（m approaeh was used to coat antibody as ligands. The resulting products were packed in columns. Series concentration of melamine was loaded on the columns to evaluate column＇s performances. UV spectrum and liquid chromatography tandem mass speetrum （LC-MS） were used to test the melamine level of eluents from columns. The mean load capacity of the prepared columns was 16.47 μg per column. Under the optimized conditions （washing condition,loading buffer,eluent eondition and regeneration buffer）,the proposed IAC has a eondensation coefficient of 66 fi）lds and it could be used over 5 times with the reeovery higher than 60%.