Suspensions of rat hepatocytes, isolated by a collagenase-based perfusion technique, were incubated with carbon tetrachloride (CCl4) for up to 60 min. Control incubated cells examined by transmission electron microscopy had an essentially normal substructure. Treatment with CCl4, however, resulted in a loss of microvilli at the cell membrane, disorganization of the RER, the appearance of condensed membranes, vacuoles within mitochondria and cellular protrusions. While some of the alterations are similar to those observed after in vivo CCl4 exposure, others are not. This suggests that the mechanism of in vitro damage may not be identical to that in vivo. Further investigation of the differences, such as the cellular protrusions, may yield increased understanding of the toxicity induced by CCl4 in the whole animal.