Livers from human fetuses between the 16th and 24th weeks of gestation were dissociated by successive dispase and collagenase digestion followed by two cycles of low-speed differential centrifugation. This improved method recovered approximately 1 X 10(7) cells (90% hepatocytes and 90% viable cells) from 4 g of liver tissue. These hepatocytes were set into primary culture and monolayer granular hepatocytes were obtained within a week. Both albumin and alpha-fetoprotein production was demonstrated in these granular hepatocytes by the immunoperoxidase method for 2 weeks and alpha-fetoprotein production in the culture medium occurred for a week by the single radial immunodiffusion method. The morphological features of the granular hepatocytes could be distinguished from those of the other type of epithelial cells with clear cytoplasm. During the cultivation period, gradual changes from granular to clear hepatocytes with high mitotic activity were found.
