A simple and efficient method for isolation of Kupffer cells

Objective To develop a simple,economical and efficient method for isolation of Kupffer cells,and to provide cells for investigating the role of Kupffer cells in liver and systematic diseases.Methods Based on the present methods,to perfusate the liver in vivo at first,then perfusate and digest the liver with type Ⅳ collagenase ex vivo,after that,the Kupffer cells were separated by density gradient centrifugation with percoll separating medium and cultured.The activity of the cells were determined by trypan blue,and the purity were assessed by Fluophot,immunochemical way and phagocytosis test.Results The cell number was about(5～10)×107 per liver,the activity were above 90%,and purity were above 95%.The isolated Kupffer cells showed polymorphism with typical polygon-like and star-like shapes,immunochemical way exhibit positive result of CD163(ED2),and the phagocytosed carbon particle were watched under microscope.Conclusion This method is simple and efficient for preparing Kupffer cells,and it doesn′t need the special equipment,so it is easy to be applied.The acquired cells with high activity and purity can be used for advanced research.