CD34^+造血干/祖细胞蛋白质组双向电泳图谱的建立

Establishment of two-dimensional gel electrophoresis profiles of proteome from CD34^+ hema- topoietic stem/progenitor cells

目的探讨骨髓(BM)和动员后外周血(MPB)CD34+细胞的蛋白质差异表达情况。方法应用免疫磁珠法分离、纯化正常人骨髓和粒细胞集落刺激因子(G-CSF)动员后外周血单个核细胞(MNCs)中的CD34+细胞,冻融法提取CD34+细胞的全细胞蛋白质进行双向电泳,并对电泳结果进行图像分析。结果纯化后CD34+细胞的平均纯度为(92.33±2.65)%;每次纯化获得的CD34+细胞数平均为(1.12±0.42)×106个。优化的双向电泳技术获得较为理想的蛋白质组图谱,显示出不同来源的CD34+细胞存在不同的蛋白质表达。结论免疫磁珠联合双向电泳适用于造血干/祖细胞的蛋白质组研究,骨髓和动员后外周血CD34+细胞的蛋白质表达存在差异。

Objective To investigate the differential expression of proteins in bone marrow (BM) and mobilized peripheral blood (MPB) CD34+ cells. Methods Immunomagnetic beads were used to separate and purify CD34+ cells from the BM and the MPB mononuclear cells (MNCs) mobilized by granulocyte colony-stimulating factor (G-CSF) of normal subjects. The whole cell proteins in CD34+ cells were extracted by freeze-thaw lysis, and applied for two-dimensional electrophoresis (2-DE), the results analyzed with image analysis software. Results The average purity of CD34+ cells was 92.33%±2.65% in output, with an average cell number of (1.12±0.42)×106. 2-DE techniques were optimized, which yielded satisfactory 2-DE profiles of the proteome, showing the different expressions of the proteins between different CD34+ cells. Conclusions Immunomagnetic beads in combination with 2-DE is applicable for research of hematopoietic stem/progenitor cells proteome. There are differences in the protein expressions between BM- and MPB-derived CD34+ cells.