Comparison of Techniques Irrigating and Reversing Rabbit Ear Vein to Digest, Isolate and Culture Endothelial Cells

Objective To establish the better method for isolating and culturing the endothelial cells (ECs). Methods The “irrigative digestion” and “reverse interiorly-exteriorly digestion” methods were performed for digesting, isolating, collecting and culturing the ear vein endothelial cells of rabbit. The trypan blue stain was used to test the cell activity. The third-passage cell was identified by factor Ⅷ related antigen. The differences between the methods in cell number, activity and purity were compared to get an optimal method. Results The number of ECs deriving from the “irrigative digestion” method had no significant difference from that deriving from the “reverse interiorly-exteriorly” method when cells were isolated from rabbit ear vein originally. However, after cultured for 5 or 10 days, the vein endothelial cells from the “irrigative digestion” method showed the growth status superior to those from the “reverse interiorly-exteriorly” method. The cultured cells had a cobble stone appearance with a strict monolayer growth, it could be observed under inverted microscope. The factor Ⅷ related antigen was tested by immunohistochemistry, it supported that the cultured cells was ECs. Conclusion The “irrigative digestion” method is the better choice to isolate endothelial cells from small vessel.