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目的:用目前常用的干细胞分离方法对正常SD大鼠的胰腺成体干细胞进行分离、培养和鉴定,为胰腺癌干细胞的进一步研究奠定基础。方法:选取正常SD大鼠胰腺用胶原酶P消化分离,并用低血清无糖培养基纯化胰腺细胞,以RT-PCR,免疫组化和Western blot方法鉴定胰腺成体干细胞标志物巢蛋白(nestin)并通过RT-PCR证明细胞中存在ABCG2(ATP-binding cassette superfamily G member2)。结果:成功分离到所需胰腺成体干细胞,并通过以上3种方法证明分离所得细胞表达巢蛋白。结论:以胶原酶P作为消化液,低血清无糖培养作为分离方法能够成功的从正常胰腺中分离得胰腺成体干细胞。
OBJECTIVE: To isolate, culture and identify adult rat pancreatic stem cells from normal SD rats by using the commonly used methods of stem cell isolation, and to lay a foundation for the further study of pancreatic cancer stem cells. Methods: The pancreatic tissues of normal SD rats were selected and digested with collagenase P, and the pancreatic cells were purified with low serum sugar-free medium. The expression of nestin in pancreatic adult stem cell markers was identified by RT-PCR, immunohistochemistry and Western blot The presence of ABCG2 (ATP-binding cassette superfamily G member2) was confirmed by RT-PCR. Results: The desired pancreatic adult stem cells were successfully isolated and the resulting cells were verified to be nestin by the above three methods. CONCLUSION: Adult rat pancreatic stem cells can be successfully isolated from normal pancreas using collagenase P as a digestive juice and low serum sugar-free culture as a separation method.