Fibroblast-like cells were isolated from liver biopsies of normal adult donors. The cells were grown in tissue culture first as a heterogeneous population, afterwards as homogeneous cultures of fibroblast-like cells. Phase contrast microscopy demonstrated that cultured human liver fibroblast-like cells grew as monolayers of slender, spindle-shaped cells in parallel arrays. By transmission electron microscopy (TEM), cultured human liver fibroblasts were seen to have morphological characteristics of in vitro fibroblasts. By immunoelectronmicroscopy, cultured fibroblast-like cells were seen to produce components of connective tissue, such as fibronectin, collagen type I, type III, and small amounts of collagen type IV. These studies demonstrate that it is possible to culture morphologically and immunologically identifiable human liver fibroblasts from normal human liver.