ABSTRACT: Previous immunohistochemical studies of the rat placenta using specific alloantisera and/or monoclonal antibodies showed that the basal zone trophoblasts stained for Pa and Aa class I major histocompatibility complex (MHC) antigens and for the human SP1-related antigen. In an effort to isolate the basal zone trophoblast cells from the rat placenta, we used these markers to assess the degree of purification of the cells separated by density gradient centrifugation using either Ficoll-Hypaque or Percoll as the gradient medium. The cells were put either on the top or at the bottom of discontinuous density gradients in the range of 1.005-1.10 or/ml. The cell separation profiles for the two media were different. With Percoll, most of the trophoblast cells (80–95%) were collected at the density gradients 1.04/1.06 and 1.06/1.08, whereas with Ficoll-Hypaque, these gradients separated only a small fraction (4–23%) of the trophoblast cells, and most of them pelleted at the bottom of the tube. The trophoblast cells separated by Ficoll-Hypaque, however, showed fewer contaminant cells than those separated by the Percoll gradients.