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目的 :探索高原环境下动物肝细胞的体外分离方法 ;方法 :采用胶原酶体外两步灌流法先进行肝叶灌流 ,再将所得肝组织悬液分别置 3 7℃温育不同时间 ,对比观测肝细胞产量、存活率、纯度及细胞接种后生长情况的差异 ;结果 :未经温育组及温育 1 0min、3 0min、60min组肝组织悬液的肝细胞产量分别为 ( 0 .85、2 .67、3 .84、4.2 9)× 1 0 6/ g湿重肝组织 ;肝细胞存活率分别为 97.6%、96.9%、88.3 %及 71 .5% ;肝细胞纯度分别为 96.2 %、95.4%、96.7%及 95.8% ;前两组细胞贴壁生长和增殖情况均较好 ,后两组则较差 ;结论 :在高原环境下先采用肝叶体外两步灌流后再行胶原酶短时间 ( 1 0min左右 )温育处理是一种既可获得较满意的细胞活性 ,同时又可显著提高细胞产量的分离肝细胞方法。
OBJECTIVE: To explore the method of in vitro isolation of animal hepatocytes in plateau environment.METHODS: Hepatic lobe perfusion was performed by collagenase in vitro two-step perfusion method, and then the resulting suspension of liver tissue was incubated at 37 ℃ for different times, Cell yield, survival rate, purity and cell growth after inoculation; Results: The liver cell yield of the liver tissue suspension in the untreated group and incubated for 10 min, 30 min and 60 min were (0.85, 2, .67,3.84,4.2 9) × 10 6 / g wet liver tissue; the survival rate of hepatocytes were 97.6%, 96.9%, 88.3% and 71.5% respectively; the purity of hepatocytes were 96.2%, 95.4 %, 96.7% and 95.8%, respectively. The adhesion and growth of cells in the first two groups were better than those in the latter two groups. Conclusion: In the plateau environment, (About 10min) incubation is a method that can achieve more satisfactory cell activity, while significantly increasing cell yield of isolated hepatocytes.