摘要
目的建立对树鼩枯否细胞(Kuffer cells,KCs)进行分离、原代培养和鉴定的方法。方法经肝活检手术分别获取6只树鼩的肝组织,后采用胶原酶体外灌注结合机械法分离肝脏细胞,经差速离心法、Percoll梯度密度离心法及贴壁法分离、纯化KCs后进行原代培养,最后通过细胞免疫组化、墨汁吞噬实验及透射电镜对KCs进行鉴定。结果树鼩肝组织分离获得的KCs数量为(1.2±0.2)×106个/g肝组织,细胞活力为90%,细胞纯度达85%以上。结论本组建立的胶原酶体外灌注结合机械分离树鼩KCs的方法稳定实用,为同一研究个体的KCs进行多次分离培养和动态观察奠定基础。
Purpose To establish a method to isolate and identify Kupffer cells for primary tissue culture from tree shrews. Methods The procedure to isolate Kupffer cells from six tree shrews includes： combine in vitro collagenase perfusion with mechanical action to separate liver cells, follow by differential speed centrifuge, Percoll density gradient eentrifugation and attachment purification. The i- dentification of Kupffer cells is performed by immunohistochemistry, ink phagocytosis experiments and transmission electron micro- scope. Results The harvest of Kupffer cell from tree shrew liver tissue were （ 1.2 ±0.2）×10^6cells/g liver, the cell viability was 90% and the purity was more than 85%. Conclusion This study established a stable and convenient method to isolate Kupffer cells from tree shrews, which mainly involved in vitro collagenase perfusion with mechanical action. This method can be applied to isolate, culture, long-term repeated observe Kupffer cells from one animal.
出处
《临床与实验病理学杂志》
CAS
CSCD
北大核心
2014年第2期159-163,共5页
Chinese Journal of Clinical and Experimental Pathology
基金
国家自然科学基金(30660171)
广西自然科学基金(桂科自0447087)
关键词
树鼩
枯否细胞
分离
原代培养
鉴定
tree shrews
Kupffer cells
isolation
primary culture
identification
