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Isolated hyperthermic perfusion of the liver was performed for 45 min in 27 pigs via hepatic artery and portal vein at mean inflow temperatures between 40.7 and 41.2°C. In two study groups B and C (n = 9 pigs each) 50 μg recombinant human tumor necrosis factor-α (rhTNFα) per kg body weight were added to the perfusate, whereas in a control group A liver perfusion was done without rhTNFα. Before reperfusion the livers were washed out with Ringer’s solution in all groups followed by a protein solution in group C. At 30 and 60 min after reperfusion the maximum systemic rhTNFα concentrations were significantly higher in group B with 68 and 61 ng/ml compared to 14.5 and 14.9 ng/ml in group C (p < 0.05). Mean systemic porcine TNFα concentration was significantly higher in group B (217 pg/ml) compared to group C (50 pg/ml) 30 min after reperfusion (p = 0.012). Survival was 7/9 in group A and C and only 2/9 in group B with 6/7 pigs dying due to severe cardiopulmonary failure within 12 h after operation. In surviving pigs of group A and C only mild and transient hepatotoxicity was registered. The presented study underlines the feasibility of high dose rhTNFα application in an isolated hyperthermic liver perfusion system. Washout of the liver with a protein solution before reperfusion reduces systemic TNFα levels as well as associated lethal cardiocirculatory and hepatotoxic side effects.