鸡囊胚细胞冷冻损伤的评价方法  

Evaluation methods of cryodamage to chicken blastodermal cells(BCs)

作　　者：王坤[1] 燕海峰[2,3] 曲湘勇[1] 何俊[1] 王庆桥 蒋隽[1] 

机构地区：[1]湖南农业大学动物科技学院,湖南长沙410128 [2]湖南省畜牧兽医研究所,湖南长沙410131 [3]湖南天心黄鸡育种有限公司,湖南长沙410143

出　　处：《中国兽医学报》2016年第6期1065-1070,共6页Chinese Journal of Veterinary Science

基　　金：国家国际科技合作专项资助项目(2013DFG31810);湖南省科技计划资助项目(2014WK2008);长沙市科技计划资助项目(K1307134-24);湖南省研究生科研创新资助项目(CX2015B279)

摘　　要：采用滤纸环法分离鸡囊胚细胞(blastodermal cells,BCs),冷冻复苏后使用二乙酰荧光素(fluorescein diacetate,FDA)测定细胞活率,以及培养24h测定细胞贴壁率简称"贴壁率法",探究2种方法在不同冷冻程序优化试验中对细胞冷冻损伤的评价效果。结果显示,FDA染色法与贴壁率法在冷冻保护剂浓度优化、冷冻及解冻速率优化试验中对细胞冷冻损伤的评价结果差异不显著(P>0.05),相关性很好,但也有一定差异;不同冷冻剂浓度优化试验中,5%DMSO冷冻保护液有细胞活率,但无贴壁现象;投入液氮温度对比试验中,-35℃以后,随着投入温度的降低,细胞活率略微升高,但贴壁率呈较明显的下降趋势。因此,2种方法均适合鸡BCs冷冻损伤评价,但贴壁率法能给出更多关于细胞状况的信息,评价结果更加真实可靠。The chicken blastodermal cells（BCs）was separated by filter paper ring from eggs,cells viability（CV）by fluorescein diacetate（FDA）,cells adherent ratio cultivated after 24h（CAR）were used to tested by evaluating effectiveness of cryodamage in different cryopreservation program.The results showed that the two methods had no significant difference（P〉0.05）for evaluating cryodamage.However,there were some differences in some cases,for example,in the experiment about optimization of DMSO concentrations,the DMSO concentration was 5%,there were cells viability,but the CAR was 0.In the experiment about freezing ratio,the CV raised slightly,but the CAR was obviously downtrend when the temperature less then-35℃.Both methods were fit for evaluating the BCs cryodamage in frozing,but the CAR could provide more information about the condition of the cells and the evaluation results were more reliable.

关 键 词：鸡 囊胚细胞 冷冻损伤 FDA荧光染色 细胞贴壁率 

分 类 号：S814.8[农业科学—畜牧学]