Isolated organ perfusion is used in clinical practice for chemotherapy in adults with malignant tumors. However, it has not been performed in children because of the size mismatch with the adult circuits. The authors have previously studied isolated liver perfusion in small animals using the self-regulating extracorporeal membrane oxygenation circuit. The present study was designed to investigate the biochemical and morphological changes in the liver during isolated liver perfusion with double bypass using automatic blood pumps. Isolated liver perfusion was performed with bypass between the hepatic and portal veins in seven weanling Yorkshire swine weighing 8.2 to 12.2 kg, at a flow rate of 20 mL/min/kg for up to 4 hours. Venous blood from the intestine and lower body was bypassed to the superior vena cava. As a result, perfusate glutamic pyruvic transaminase and lactate concentrations did not change during liver perfusion. On gross inspection, the surface of the liver was mottled. Microscopically, normal histology of the hepatic parenchyma and portal tract structures was preserved. Transmission electron microscopy showed no gross structural abnormalities in most of the hepatocytes for up to 4 hours. However, swelling of the mitochondria and smooth endoplasmic reticulum was seen occasionally in a very small number of the hepatocytes after more than 3 hours of perfusion. Glycogen granules decreased with time in some animals. Isolated liver perfusion at 20 mL/min/kg of perfusion flow can be performed safely for up to 4 hours with nearly intact hepatocellular function and morphology.