人脐带血有核细胞冷冻干燥保存实验初步研究

RESEARCH ON FREEZE-DRYING OF THE NUCLEATED CELL OF HUMAN CORD BLOOD

脐带血中的造血干细胞有着广泛的应用前景 ,随之而产生的保存问题已成为关注的热点。目前已有低温保存的研究 ,但未有冻干成功的报道。实验试图用冻干的方法来长期保存脐带血中的造血干细胞。冻干过程中选用PVP、HES和各种糖类作为保护剂 ,一次干燥阶段搁板温度控制在 - 30℃ ,二次干燥时搁板温度控制在 +1 5℃ ,整个冻干过程为 52 .5h。冻干后复水性能极佳 ,能够在 30s内完全复水 ,用显微染色观察 ,细胞形态完整 ;用流式细胞仪进行PI染色和CD34+抗体跟踪检测 ,测得有核细胞存活率为 55 .67% ,CD34+ 抗体跟踪检测得到CD34+ 细胞占淋巴细胞的 3 .61 %。

The research on hematogenic stem cells of cord blood has become more popular. How to preserve the cord blood stem cells have been researched by many researchers. At present, cord blood is preserved in ultra low temperatures. In this research, it was preserved it by freeze drying. Firstly, the samples with different concentrations of PVP,HES and saccharides were frozen to -40℃ on the lyophilizer's shelf. Afterwards, they were vacuum dried at selected shelf temperature at -30℃ for main drying stage, and then at 15℃ for second drying, the entire time of the freeze drying is 52.5hs. The dried samples were resuspended in an isotonic phosphate buffered saline solution for rehydration and tested by microscope and flow cytometer. The protocol of 40%PVP+20%sucrose had the best result. Flow Cytometer (FCM) measured the livability of the nucleated cells,and it was 55.67%. The ratio of the number of CD34 + to lymphocytes was 3.61%.