免疫磁珠纯化小鼠精原干细胞的研究

精原干细胞（spermatogonial stem cells，SSCs）富集纯化是利用SSCs进行基因修饰新方法等研究的前提基础。采用免疫磁珠分选法，使用干细胞抗体CD90.2进行小鼠SSCs的纯化富集，并采用流式细胞分析法和定量PCR验证了磁珠分选效率。流式细胞分析结果：免疫磁珠分选后SSCs纯度为50.11%。荧光定量PCR检测结果：磁珠分选后支持细胞特异表达基因 GATA4 显著下调（6倍）、SSCs表达基因 GFRα-1 上调（6.5倍）、生殖干细胞特异表达基因 OCT4 极显著上调（5.9倍），3个基因相对表达量的变化说明，免疫磁珠分选效率为6倍。流式细胞分析法所产生的偏差可能是受到了未解离磁珠及SSCs本身转基因荧光的影响。

Research on Rurification of Mouse Spermatogonial Stem Cells Using Magnetic Microbeads

Enrichment and purification of spermatogonial stem cells (SSCs) is a prerequisite for genetic modification methods by SSCs. Stem cells antibody CD90.2 was used to enrich and purify mice SSCs through magnetic activated cell sorting(MACS) method, flow cytometry analysis(FACS) and Real-Time Polymerase Chain Reaction (RT-PCR) were used to detect the sorting efficiency. The sorting result using FACS showed that the purity of SSCs was 50.11% after sorting. However, the data by RT-PCR suggested that the expression of GATA4, a special-expression gene of sertoli cells, was downregulated about 6 times, and the expression of GFRα-1 and OCT4, two special-expression genes of germ stem cells, was upregulated 6.5 and 5.9 times respectively after-sorting. The relative expression of those three genes indicated that the enrichment efficiency using MACS was 6 times. The deviation between FACS and RT-PCR may come from undissociated magnetic bead and genetically modified fluorescence of SSCs.