Comparison of four 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 2-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.
Gerlach
J C
JC
Surgical Department, Universitätsklinikum Rudolf Virchow, Freie Universität Berlin, Germany.
Brombacher
J
J
Klöppel
K
K
Schnoy
N
N
Neuhaus
P
P
D000818:Animals; D002469:Cell Separation / Q000379:methods*; D002470:Cell Survival; D017364:Collagenases; D004492:Edetic Acid; D006801:Humans; D008099:Liver / Q000166:cytology*; D008297:Male; D010477:Perfusion; D013552:Swine
D004492:Edetic Acid; D017364:Collagenases