高效亲和膜色谱快速分析及小量制备蛋白质

Fast Assay and Mini-preparation of Protein by High Performance Membrane Affinity Chromatography

以甲基丙烯酸缩水甘油酯纤维素复合膜为基质，分别以蛋白Ａ（ＰｒｏｔｅｉｎＡ）、人免疫球蛋白Ｇ（ＨＩｇＧ）、三嗪染料（ＣｉｂａｃｒｏｎｂｌｕｅＦ３ＧＡ）、亚胺二乙酸铜离子为配基，用不同方法制备了适合于分析及小量制备的高效亲和膜色谱介质，并对高效亲和柱的基本性能及其应用于各种相应蛋白的定量测定情况进行了考察。利用这种方法可以针对不同的目标蛋白及所存在的环境采用不同的配基，对各种蛋白的定量测定及小量制备可达到较为满意的结果。

Several novel affinity ligands,protein A,human immunoglobulin G,iminodiacetate (IDA) Cu(Ⅱ) and cibacron blue F3GA have been coupled to GMA modified cellulose membrane matrices for rapid assay of protein in high performance membrane affinity chromatography(HPMAC).The non specific adsorption of the high performance membrane media has been studied by injecting BSA on the protein A column.The media without arm has no non specific absorbance of BSA.The calibration curve showed a good linearity (curvefit coefficient >0 997) for all sorts of media.The total time for separation and determination was within 5 min when flow rate was 1 ml/min,the total time of rapid assay was within 30s when flow rate was 3ml/min.Relative standard deviation of peak areas was less than 3 2% for the proteins in standard solution and human plasma.The media could also be used for mini preparation of protein.The most HIgG that could be specifically adsorbed once was 1.2 mg and 0 19 mg by the protein A column with hexyldiamine as the arm and that without the arm respectively.The column with the arm could also maximally non specifically adsorb 0 023 mg BSA.