论文部分内容阅读
目的针对人血清双向电泳流程中样品制备的不同方法,本研究进行了系统的比较和分析。方法采用丙酮沉淀法,三氯醋酸沉淀法,三氯醋酸/丙酮沉淀法,蛋白纯化试剂盒,去白蛋白及免疫球蛋白试剂盒预处理血清样品;双向电泳分离;硝酸银染色;软件分析所得蛋白质斑点。结果对丙酮沉淀法,三氯醋酸沉淀法,三氯醋酸/丙酮沉淀法,蛋白纯化试剂盒处理血清所得到的2-DE图谱统计分析,得到平均蛋白质点数分别为1 802±12,1 648±10,1 727±13,1 670±14;对去除白蛋白及免疫球蛋白血清所得到的双向电泳(2-DE)图谱与丙酮沉淀血清所得到的图谱进行统计,得到平均蛋白质点数分别为1 212±16,1 258±12,并对其中差异蛋白进行了质谱鉴定。结论结果表明不同血清沉淀方法对血清的双向电泳各有优劣;血清去除白蛋白及免疫球蛋白之后,可以提高一些低丰度蛋白的检出,但会导致部分蛋白的丢失。
Aim To compare the different methods of sample preparation in human serum two-dimensional electrophoresis flow, this study systematically compared and analyzed. Methods Serum samples were pretreated with acetone precipitation, trichloroacetic acid precipitation, trichloroacetic acid / acetone precipitation, protein purification kit, albumin removal and immunoglobulin kit; two-dimensional electrophoresis separation; silver nitrate staining; Protein spots. Results The statistical analysis of the 2-DE patterns obtained from the acetone precipitation, trichloroacetic acid precipitation, trichloroacetic acid / acetone precipitation, and protein purification kits resulted in average protein spots of 1 802 ± 12 and 1 648 ± 10,1 727 ± 13,1 670 ± 14; The results of two-dimensional gel electrophoresis (2-DE) and serum albumin obtained by removing albumin and immunoglobulin serum were statistically analyzed. The average number of protein spots was 1 212 ± 16,1 258 ± 12, and the differential proteins were identified by mass spectrometry. Conclusions The results show that different serum sedimentation methods have the advantages and disadvantages of two-dimensional electrophoresis. Serum removal of albumin and immunoglobulin can increase the detection of some low-abundance proteins but lead to the loss of some proteins.