基于自组装多肽的无血清细胞冷冻保存  

Serum⁃free cell cryopreservation based on self⁃assembled peptides

作　　者：郑义哲 李东东 王蕾 张萱 李艳宏 刘坤 王博 阎少多 付秋霞 梁俊 ZHENG Yizhe;LI Dongdong;WANG Lei;ZHANG Xuan;LI Yanhong;LIU Kun;WANG Bo;YAN Shaoduo;FU Qiuxia;LIANG Jun(Department of Light Industry Science and Engineering,Tianjin University of Science and Technology,Tianjin 300457,China;不详)

机构地区：[1]天津科技大学轻工科学与工程学院,天津300457 [2]军事医学研究院卫生勤务与血液研究所,北京100039

出　　处：《中国生物制品学杂志》2024年第5期605-610,616,共7页Chinese Journal of Biologicals

基　　金：国家重点研发计划(2021YFC2101400).

摘　　要：目的实现基于自组装多肽(Peptide)的无血清低DMSO的高效细胞冷冻保存。方法使用胎牛血清(FBS)、DMSO和Peptide 3种冷冻保护剂(cryoprotectant,CPA)进行组合冷冻保存Hepa1-6细胞。采用共聚焦显微成像、流式细胞术及细胞计数等方法检测冻存后细胞的存活率、回收率及增殖能力,并通过差示扫描量热(differential scanning calorimetry,DSC)法检测CPA抑冰性能,最终确定一种用于细胞冻存的高效CPA。结果Peptide可在DMSO溶液中形成纤维网络水凝胶封装细胞,支撑细胞呈三维(3D)立体分布。在无血清条件下,与5%DMSO及0.1wt%Peptide组相比,用0.1wt%Peptide联合5%DMSO作为新型复合CPA冻存Hepa1-6细胞,可显著提高细胞存活率及冻存后回收率,且与10%DMSO+20%FBS组相比差异无统计学意义(t分别为0.983和2.164,P均>0.05)。此外,与10%DMSO+20%FBS组相比,0.1wt%Peptide+5%DMSO组冻融后细胞的增殖能力差异也无统计学意义(t=3.513,P>0.05)。DSC测试显示,基于Peptide的CPA抑冰性能良好。结论使用0.1wt%Peptide联合5%DMSO作为新型复合CPA具有良好的细胞冻存效果。Objective To achieve serum-free and low-DMSO efficient cell cryopreservation based on self-assembled peptides(Peptide).Methods Hepa1-6 cells were cryopreserved by combining three cryoprotectants(CPAs),FBS,DMSO and Peptide.Confocal microscopy,flow cytometry and cell counting were used to measure the survival rate,recovery rate and proliferation ability of the cells after freezing,and the ice inhibition of CPAs was tested by differential scanning calorimetry(DSC).Finally,a CPA with high efficiency for cell cryopreservation was determined.Results Peptide formed a fibrous network hydrogel in DMSO solution to encapsulate cells and supported them in a three-dimensional(3D)stereological distribution.Under serum-free conditions,the use of 0.1 wt%Peptide combined with 5%DMSO as a novel composite CPA for cryopreservation of Hepa1-6 cells significantly improved the survival rate and post-freezing recovery rate compared with the 5%DMSO and 0.1 wt%Peptide groups,and there was no significant difference with the 10%DMSO+20%FBS group(t=0.983 and 2.164,respectively,each P>0.05).In addition,compared with the 10%DMSO+20%FBS group,0.1 wt%Peptide+5%DMSO group showed no significant difference in cell proliferation ability after freezing-thawing(t=3.513,P>0.05).In the end,DSC tests showed that Peptide-based CPA had good ice inhibition performance.Conclusion The use of 0.1 wt%Peptide combined with 5%DMSO as a novel composite CPA has a good effect on cell cryopreservation.

关 键 词：细胞冻存 自组装多肽 无血清 DMSO 

分 类 号：Q819[生物学—生物工程] Q-33