骨髓白血病细胞蛋白质组双向电泳技术的建立与优化

Establishment and Optimization of Two-dimensional Gel Electrophoresis Technique of Proteome from Bone Marrow Leukemia Cells

本研究旨在建立和优化人骨髓白血病细胞蛋白质组双向电泳分析方法,以获得分辨率高、重复性好的图谱。提取骨髓中白血病细胞总蛋白质,以等电聚焦为第一向和垂直SDS-PAGE为第二向分离蛋白,显色后用PDQuest 7.4软件分析电泳图谱,比较不同的实验条件对图谱的影响。结果显示:通过优化样品制备和电泳中的实验环节,获得了蛋白点分离清晰的图谱,平均蛋白点数为780±73,重复实验所得蛋白点匹配率为82±5%。结论:成功建立了人骨髓白血病细胞的蛋白质组双向电泳分析方法,该方法适用于后续各亚型白血病比较蛋白组学研究。

The aim of this study was to establish and optimize two-dimensional electrophoresis method for human bone marrow leukemia cells in order to obtain the profiles with high resolution and reproducibility. The total protein was extracted and separated by isoelectric focusing （IEF） and SDS polyacrylamide gel electrophoresis （SDS-PAGE）. The gels were stained with silver nitrate or Coomassie brilliant blue, and then scanned and analyzed with PDQuest 7.4 analysis software. The effects of different protein preparation methods and electrophoresis conditions on the profiles were compared. The results indicated that by optimizing preparation of protein sample and electrophoresis protocols, clear profiles with 780 ± 73 well separated protein spots on an average were obtained and the match rate was 82 ± 5% between reproducible gels from leukemia cells of different sub-type. It is concluded that the two-dimensional electrophoresis method of proteome from human bone marrow leukemia cells is established successfully and is suitable for the further comparative proteomic research between leukemia of different types.