摘要
采用两步酶解和差异贴壁法分离、纯化鸡精原干细胞(SSCs),比较了6种冷冻保护液对其冷冻保存效果,结果表明:FBS浓度为10%时,10%DMSO与10%甘油解冻后细胞存活率分别为54.05%和37.49%,差异显著(P<0.05);以10%DMSO为冷冻保护剂时,FBS浓度从10%增加到20%,解冻后细胞存活率分别为54.05%和69.06%,差异显著(P<0.05);在冷冻液Ⅰ基础上,添加3种不同浓度的蔗糖溶液,解冻后细胞存活率分别为52.49%、47.65%和51.94%,三者之间差异不显著(P>0.05),且与冷冻液Ⅰ组差异也不显著(P>0.05);将解冻后细胞接种饲养层上培养,除甘油组外,SSCs均能增殖并形成AKP阳性集落,但10%DMSO加上20%FBS组细胞增殖速度和集落数优于其他组合,表明10%DMSO加上20%FBS为鸡SSCs最佳冷冻保护剂。
Compare the freezing effect of six different concentration cryopreservation medium on the chicken spermatogonial stem ceils （SSCs） which were isolated using collagenase and trypsin two-step enzymatic digestion, and purified by differential adhesion. The results showed that cells frozen/thawed in FBS-based medium （DMEM） containing 10% DMSO had a significantly better cell survival rate than 10% glycerol, and the cell survival rate was 54.05% and 37.49%, respectively （P 〈 0.05）; An increase in the concentration of FBS in the DMSO-based medium to 20% had a better survival rate, and the cell survival rate was 54.05% and 69.06%, respectively（P 〈 0.05 ）; Furthermore, inclusion of three different sucrose concentration in FBS-based medium containing 10% DMSO resulted in no improvement of cell survival （P 〉 0.05）, and the three sucrose concentration cryopreservation medium had no significant difference among themselves （P 〉 0.05）; When the thawed cells were cultured on feeder layers, with the exception of glycerol group, SSCs can proliferate and form AKP-positive colony, but the cell proliferation rate and the number of colony with medium containing 10% DMSO and 20% FBS was better than other combinations, showing that cryoservation in DMEM-based medium containing 10% DMSO and 20% FBS using fast freezing protocol was an effective way to cryopreserve chicken SSCs.
出处
《中国畜牧杂志》
CAS
北大核心
2009年第21期16-19,共4页
Chinese Journal of Animal Science
基金
国家自然科学基金项目(30500270)
浙江省教育厅科研项目(20070479)
关键词
精原干细胞
冷冻保存
鸡
spermatogonial stem cells
cryopreservation
chicken
