COMPARISON OF 4 METHODS FOR MASS HEPATOCYTE ISOLATION FROM PIG AND HUMAN LIVERS

Using the pig liver, parameters for large scale hepatocyte isolation were studied in order to develop a technique suitable for human organs. These investigations led to a 5-step modification of the original 2-step method. Four groups were compared. A nonenzymatic EDTA perfusion technique has been shown to be inconvenient for mass cell isolation. The enzymatic a-step perfusion, using 0.08% collagenase and 20-kg pigs, resulted in a mean hepatocyte viability of 61+/- 1.9%, with a mean yield of 67+/-6.5% wet weight of the organ. The enzymatic 5-step method resulted in a mean hepatocyte viability of 74+/-1.7% with a mean yield of 80+/-1.8% wet weight. Five-step portal venous perfusion in combination with arterial perfusion resulted in 76+/-2.6% viability with a yield of 82+/-6.1%. The results were dependent on collagenase concentration and weight of the donors, improving with decreasing body weight. The 5-step method with combined arterial and portal vein perfusion developed for pig liver was used for mass human liver cell isolation with a minimum viability of 57% and a minimum yield of 58% wet weight.