We have investigated some of the physical properties of mouse spleen cells involved in the mixed leukocyte reaction (MLR) and in graft-vs-host disease (GVHD). When “transformed” cells were separated by velocity sedimentation they were found to be large cells sedimenting at 6.1 mm/hr. On density centrifugation, these cells are found in the light density region of the gradient (modal density, 1.055 g/cm3). In contrast, --positive cells and the precursors of “transformed” cells, i.e., cells capable of responding in the MLR and to phytohemagglutinin (PHA), are found in a higher density region (modal density, 1.071 g/cm3). Transplantation of cells from this region into appropriate recipients produces death from acute GVHD. However, cells from regions deficient in MLR- and PHA-responsive cells, but rich in hematopoietic colony-forming cells (modal density, 1.060 g/cm3), do not produce acute GVHD. Therefore, the density characteristics of ϑ-positive cells and of cells responding in the MLR and to PHA are similar to those of cells producing acute GVHD. This result is in keeping with the possibility that these classes of cells are either identical or subpopulations of a relatively homogeneous single population of cells.