Current bioartificial liver devices are based on the use of a large mass of hepatocytes exhibiting differentiated metabolic function. The pig has become a source of interest for the acquisition of such cells— however, harvesting a large mass of highly viable cells has met with difficulty. This study describes a technique for harvesting large quantities of hepatocytes at viabilities greater than 90% and also describes several features documenting differentiated function. Pigs, 6 to 10 kg body weight, underwent in situ two-step whole liver perfusion (ethylene glycol tetraacetic acid and collagenase) and ex vivo cell harvest. Harvests yielded an average of 19.5 billion cells with an average viability of 94.6%. Hepatocytes were then entrapped in type I collagen (3×105cells/well) and cultured in serum-free media for 5 days. Pig hepatocytes produced stable amounts of albumin and maintained cytochrome P-450 and glucuronidation activity over 5 days, as shown by the metabolism of lidocaine and 4-methylumbelliferone. These data indicate that pig hepatocytes can be harvested with high yields and can retain viability and differentiated function over at least 5 days of culture, and therefore should prove to be an excellent source of hepatocytes for bioartificial liver devices.