Abstract: In order to screen appropriate methods for the isolation and culture of the skin fibroblasts from Guangxi Bama mini-pig, five enzyme digestion methods and a tissue culture method were compared in the isolation of fibroblasts of skins from different parts of mini-pig including ear, leg, abdomen, back, and tail . The results indicated that the density of fibroblasts isolated with enzyme digestion methodⅠwas significantly higher than that in other treatment groups (P<0.05), and there was no significant difference in the cell death rates between fibroblasts from the shin treated with enzyme digestion methodⅠand that with enzyme digestion method Ⅳ (P>0.05), but the cell death rates in those two treatment groups were significantly lower than that in other groups, respectively (P<0.05). Fibroblast could be isolated from the thawed ear shin tissues after frozen for 10 days with enzyme digestion methodⅠ, but the density of fibroblasts and their survival rate were significantly lower than that from the fresh ear shin tissues with the same method (p<0.01). The cell confluency rate of fibroblasts would not reach to about 100% until being cultured for 12 days. Compared to other parts of mini-pig, ear skin was more suitable to isolate the fibroblasts, because the density of fibroblasts isolated from ear skin tissues was significantly higher than that from other parts of mini-pig (P<0.05), and the attachment and growth of those fibroblasts were very well. Ear skin fibroblasts were able be passaged up to more than passage 10, but the viability of cells at passage 3 to 4 was higher than that of cells at other passage. The cell growth curve of ear skin fibroblasts at passage 3 was a typical "S" shape. Therefore, it is better to isolate the fibroblasts from the ear skin of Guangxi Bama mini-pig with the digestion of 0.25% trypsase and 0.04% EDTA for 30 min, follow by 0.2% collagenase for 30 min.

Key words: Guangxi Bama mini-pig, skin fibroblasts, enzyme digestion method, tissue culture method