华北农学报2011，Vol.26Issue(z1)：206-212,7.
鲫鱼肝细胞分离与原代培养方法的优化
Optimization of Isolation and Primary Culture of Hepatocytes in Crucian Carp (Carassius auratus)
摘要
Abstract
In order to establish a stable primary culture model of hepatocytes in Carassius auratus,3 different I solation methods (tissue culture,mechanical speration and pancreatin digestion) and culture conditions of primary hepatocytes culture were compared in the present study. The survival rate was tested with trypan blue exclusion and the viability was assessed with WST-1 ;the morphological changes of cells in long-term culture were also observed. The results showed that in tissue culture, hepatocytes migrated from liver and proliferated after 4-5 days, compare with the other methods,it took much longer time for the hepatocytes to move out and it was difficult to collect the cells for further culture. The number and viability of the hepatocytes obtained by mechanical separation were obvi ously lower than by pancreatin digestion. The liver tissue digested by 0. 1% trypsin for 30 min yielded 1. 47 × 108 cell/g (liver weight) and the viability was 91. 43%. Both culture medium and concentration of serum influenced the attachment and proliferation of the hepatocytes. The cell cultivated in M199 basal medium supplemented with 10% fetal calf serum at 28℃ ,4% CO2 for 8 -9 days can sub-cultured.关键词
鲫鱼/肝细胞/胰蛋白酶/原代培养Key words
Carassius auratus/ Hepatocytes/ Trypsin/ Primary culture分类
农业科技引用本文复制引用
贾睿,曹丽萍,丁炜东,杜金梁,徐跑,殷国俊..鲫鱼肝细胞分离与原代培养方法的优化[J].华北农学报,2011,26(z1):206-212,7.基金项目
科技部国际科技合作项目(2009DFA32620) （2009DFA32620）
无锡市科技计划(国际科技合作)(CZE00906) （国际科技合作）
中央级公益性科研院所基本科研业务费专项基金(2011JBFC05) （2011JBFC05）
