不同冷冻保护剂和冷冻方法对小鼠耳皮肤成纤维细胞冻存效果的影响  

Effects of Different Cryoprotectants and Methods on Cryopreservation of Mouse Ear Skin Fibroblasts

作　　者：肖雄[1] 邓玉金[1] 赵海龙[1] 吴有博[1] 李跃民[1] 

机构地区：[1]西南大学动物科技学院,重庆400715

出　　处：《动物医学进展》2012年第5期69-74,共6页Progress In Veterinary Medicine

基　　金：重庆市自然科学基金计划项目(CSTC;2009BB5301);西南大学动物科技学院青年教师科研基金项目

摘　　要：通过比较不同冷冻保存方法和冷冻保护剂对小鼠耳皮肤成纤维细胞冷冻-解冻复苏后细胞存活率、48h贴壁率和细胞生长曲线的影响,筛选适宜的小鼠耳皮肤成纤维细胞冷冻保存方法和冷冻保护剂。结果表明,以100mL/L二甲基亚砜(DMSO)作为冷冻保护剂进行小鼠耳皮肤成纤维细胞冷冻保存时,方法3处理组解冻复苏后细胞存活率和培养48h细胞贴壁率均高于方法2处理组(P>0.05),并分别极显著高于方法1和方法4。采用方法3进行冷冻保存时,以100mL/L DMSO作为冷冻剂的细胞贴壁率显著高于100mL/L甘油(GL)组(P<0.05),且冷冻解冻后的细胞呈现正常的分裂增殖生长模式。因此,宜选择100mL/L胎牛血清(FBS)+100mL/L DMSO+DMEM作为冷冻保护液,采用方法3进行小鼠耳皮肤成纤维细胞的冷冻保存。To explore the appropriate cryopreservation system for mouse ear skin fibroblasts,the effects of different cryopreservation methods and cryoprotectants on survival rate and adhesive rate of mouse ear skin fibroblasts after being revived were studied.The results showed that when 100mL / L dimethyl sulfoxide（DMSO） was used as the cryoprotectant,after mouse ear skin fibroblasts were cryopreservated with method 3,i.e.mouse ear skin fibroblasts were precooled and balanced for 0.5hat 4℃,the freezing tube were hanged in gaseous nitrogen for 4hand then were sunk into liquid nitrogen and thawed,the survival rate and adherence rate of cells cultured for 48hwere higher than those with method 2,i.e.mouse ear skin fibroblasts were precooled and balanced for 0.5hat 4℃,then cryopreservated for 4hat-20℃and cryopreservated for 48hat-80℃,finally the freezing tube were sunk into liquid nitrogen.（P〈0.05）,method 1,i.e.mouse ear skin fibroblasts were precooled and balanced for 2hat 4℃,then cryopreservated for 4hat-20℃,finally the freezing tube were sunk into liquid nitrogen（P〈0.01） and method 4（P〈0.01） i.e.the freezing tube were sunk into liquid nitrogen respectively.When mouse ear skin fibroblasts cryopreservated in DMEM supplemented with 100mL / L fetal bovine serum（FBS） and 100mL / L DMSO with method 3,the adherence rate of cells cultured for 48hwas significantly higher than those with 100mL / L FBS and 100mL / L glycerol（GL）（P〈0.05） and proliferation of the former was normal.Therefore,cryopreservation of mouse ear skin fibroblasts can be improved by being cryopreservated in DMEM supplemented with 100mL / L FBS and 100mL / L DMSO with method 3.

关 键 词：冷冻保存 解冻 耳皮肤成纤维细胞 小鼠 

分 类 号：S852.165[农业科学—基础兽医学]