猪皮肤组织块冷冻保存方法的研究

Study on cryopreservation methods for skin tissue of pig

以二甲基亚砜、甘油和乙二醇为冷冻保护剂,以3种浓度(10%、20%、30%)在2种降温条件下探讨适合猪耳皮肤组织冷冻保存的最适条件和方法以及复苏后对皮肤组织生长能力的影响。结果表明:快速冷冻条件下各处理均无组织块生长。慢速冷冻条件下复苏后组织块的存活率均为100%,各冷冻保护液3种浓度的平均生长率分别为14.29%、56.52%和8.70%,差异显著。20%甘油保护液下采用慢速冷冻方法复苏后组织块的生长率达82.35%,极显著高于其他组和对照组。新鲜和冻存后的细胞培养均采用组织块培养法,EDTA-牛血浆贴覆,97.78%的新鲜样本组织块能较好生长,且扩展能力强。

The aim of the study was to find the most efficient cryopreservation method for skin tissue of swine ear,and to explore a way and system for producing high-proliferated cell population.We compared a variety of cooling conditions to investigate the effect of freezing on the viability of skin tissue through different freezing methods,different cryoprotectants such as DMSO,GLY and EG,in three different cryoprotectant concentrations（10%,20%,30%）.The viability of the frozen-thawed ear tissue was examined by Hoechst fluocences staining after culturing 18 d in vitro.The results showed that： none of the tissue proliferated under quick-freezing,however,100% of tissue survived under slow-freezing,with average growth rate（14.29%,56.52%,8.70%） in three concentrations of different cryoprotectants respectively.The difference was significant.After all,the highest growth rate was 82.35% with 20% glycerine protected under slow-freezing,which was significantly higher than other group and the control group.The skin fibroblasts were cultured by tissue pieces,with EDTA-blood plasma adhering.The results showed equated 97.78% fresh tissues had grown,and they could proliferate powerfully.