APA-3T3细胞微囊的深低温保存

Method of cytoperservation APA microencapsulated 3T3 cells

目的:通过优选冻存液的成份和浓度,建立低温保存APA微囊化小鼠成纤维细胞(3T3细胞)的方法。方法:用海藻酸钙-多聚赖氨酸-海藻酸钙微囊(APA)包裹3T3细胞制成APA-3T3细胞微囊。以高糖DMEM培养液为基础冻存液,分别加入不同浓度的二甲基亚砜(DMSO)或牛血清白蛋白(BSA)。分别将不同的冻存液加入APA-3T3中。按程序控制降温梯度,置于液氮中保存。快速复温后,光镜下观察APA-3T3的形态、细胞活率及膜通透性。结果:①10%DMSO组的微囊内细胞活率降低最少,细胞活率达(75.68±1.54)%,P<0.01。微囊完好率达到(94.48±0.90)%,P<0.01。②与其它组相比,4%BSA组的微囊内细胞活率降低程度最小,活率达(73±1.26)%,P<0.01。微囊完好率最高,为(85.1±0.82)%,P<0.01。③加入不同成份和浓度的冻存液对微囊的通透性没有明显影响。结论:在冻存液中加入10%DMSO和4%BSA,可在低温保存的过程中较好地保护细胞的存活及微囊的完整性。

Objective：To compare the effects of frozen stock solutions with different components and concentration on low temperature preservation of APA microencapsulated mouse fibroblast cells （ APA-3T3）, and to explore the optimal condition of APA microencapsulated cells low temperature preservation. Methods： Capsulated 3T3 cells with Alginate-Polylysine-Alginate （APA） to get APA-3T3, and programmed reduced the temperature of APA-3T3 in frozen stock solutions with different concentration of DMSO and BSA, and then preserved in liquid nitrogen. Checked the form and permeability of APA-3T3 after rapid rewarming, and recorded the motility rate of cells with trypan blue dyeing. Results：Compared with other DMSO groups, the 10% DMSO group has the maximal motility rate, about （75.68 ± 1.54） %, P 〈 0.01, and microcapsule undamaged rate, about （94.48 ± 0.90）%, P 〈 0.01; Compared with other BSA groups, the 4% BSA group has the maximal motility rate, about（73 ± 1.26） %, P 〈0.01, and microcapsule undamaged rate, about（85.1±0.82） %, P 〈0.01 ; The frozen stock solutions with different components and concentration have no remarkable effect on the permeability of microcapsule. Conclusion： The frozen stock solution with 10% DMSO and 4% BSA can preserve the cells and permeability of microcapsules in the process of low temperature preservation.