冻干外周血单个核细胞制备及性能分析

Preparation and characterization of lyophilized peripheral blood mononuclear cells

目的:通过研究冷冻干燥(冻干)前后人外周血单个核细胞(PBMCs)特性变化,分析冻干对PBMCs的影响。研究不同保存条件下冻干后样品的性能变化,为研制可长期保存的细胞制品提供可能的解决方案。方法:从人全血中分离PBMCs,在等渗固定液中固定后,通过不同的工艺冻干,以流式细胞术检测细胞大小、胞内颗粒多少、特异性抗原表达和平均荧光强度等特性。结果:冻干不改变细胞大小、胞内颗粒多少和CD3/CD4/CD8阳性细胞百分率。同一组细胞冻干前后CD3^+细胞阳性率分别为76.23%±0.68%和76.84%±1.5%;CD4^+细胞阳性率分别为41.22%±0.15%和40.41%±0.77%;CD8+细胞阳性率分别为29.05%±0.2%和28.70%±0.54%。在6个月的研究周期内,-20℃和2~8℃条件下保存的样本,细胞大小、胞内颗粒多少和阳性细胞百分率未见改变。20~25℃和37℃保存的样本,14 d内均未见以上特性改变。结论:冻干不改变细胞大小,胞内颗粒多少及其所表达的白细胞分化抗原数量等特性。并且冻干细胞在-20℃或者2~8℃条件下稳定,可长期保存。

Objective:To analyze the influence of freeze-drying on peripheral blood mononuclear mononuclear cell by studying the characteristic changes of PBMCs before and after freeze-drying.To study the properties of freezedried samples under different storage conditions,and to find a possible method to make long-term preserved cell products.Methods:PBMCs were separated from human whole blood and fixed in the isotonic buffer before freeze-dried by different processes.The characteristics of the PBMCs such as cell size,granularity,cell subtypes and average fluorescence intensity(MFI)were analyzed with flow cytometry.Results:There were no effects on cell size,granularity and cell subtype with processes of lyophilization.The percentage of CD3^+cells were 76.23%±0.68%and 76.84%±1.5%;The percentage of CD4^+cells were 41.22%±0.25%and 40.41%±0.77%;The percentage of CD8+cells were 29.05%±0.2%and 28.70%±0.54%before and after lyophilization respectively.The cell size,granulanty and subtypes of PBMCs were not changed after lyophilization.Conclusion:The cell size,granularity and subtypes of PBMCs were not changed after lyophilization,and freezedried cells were stable at-20℃and 2-8℃for long storage.