[Mixed enzyme applied to develop the method on BAlb/c mouse Kupffer cell isolated and cultured in vitro].
OBJECTIVE
To research the reliable method for the isolation and culture of Kupffer cell in BALB/c mouse by mixed enzyme.


METHODS
Kupffer cells were isolated from liver by in situ perfusion and digestion with 0.1% IV collagenase, 0.2% pronase and 0.01% Dnase I, and by percoll density gradient centrifugation. Kupffer cells were identified by fluorescence microscope, immunohistochemistry and cell endocytosis effect.


RESULTS
Kupffer cells were isolated successfully with high purity, the yield of (2-3) 10(6)/per mouse liver and the identification that 0.4% trypan blue indicated that the cells survival rate and purity were more than 96% and more than 92% respectively. The shape of Kupffer cell appeared to be multiplicity, irregularity, polygon and multiangular. Kupffer cells showed lysozyme positive by immunohistochemistry staining. And particles of India ink were found in cytoplasm.


CONCLUSIONS
Here described technique for isolation and culture of Kupffer cells is simple and reliable, and can be used for preparing Kupffer cells with high yield, activity and purity.
Zheng
Xue-Lian
XL
Department of Digestive Surgery, West China Hospital, Sichuan University, Chengdu 610041, China.
Yan
Mao-Lin
ML
Liao
Da-Qing
DQ
Zhou
Zong-Guang
ZG
Chen
Ke-Ling
KL
D000818:Animals; D002469:Cell Separation / Q000379:methods*; D048430:Cell Shape; D002470:Cell Survival; D002478:Cells, Cultured; D017364:Collagenases / Q000378:metabolism; D007150:Immunohistochemistry; D007728:Kupffer Cells / Q000166:cytology* / Q000201:enzymology; D008099:Liver / Q000166:cytology; D051379:Mice; D008807:Mice, Inbred BALB C; D009113:Muramidase / Q000378:metabolism; D011402:Pronase / Q000378:metabolism; D015203:Reproducibility of Results
D009113:Muramidase; D017364:Collagenases; D011402:Pronase