Direct binding of 125I-iodinated anti-H-2Kb monoclonal antibody (B8-24-3) to EL4 cells indicated a similar association constant but a 2.7-fold higher H-2Kb antigenic density when the EL4 cells were grown in ascites (2.0 X 10(5) sites/cell) than in tissue culture (7.5 X 10(4) sites/cell). The membrane fluidity of the isolated plasma membrane fractions, measured by using a membrane-sensitive spin label, was essentially identical in the two differently cultured EL4 cells. There were some differences in the cytoskeletal proteins that were isolated from the plasma membrane fractions of the EL4 lines and analyzed by two-dimensional gel electrophoresis. These differences in the H-2Kb antigenic density and the cytoskeletal proteins may contribute to the 2.3-fold higher Vmax and the increased binding of the EL4 ascites target cells to allogeneically primed anti-H-2Kb cytotoxic T cells.