Experiments have been carried out to assess the immunostimulatory activity of a hamster IgM mAb (mAb5D3) with specificity for an 80-kDa LPS-binding protein expressed on murine macrophages and monocytes. The addition of mAb5D3 to cultures of murine bone marrow-derived macrophages activated these cells to become tumoricidal for mastocytoma cells in vitro. The activity of mAb5D3 was enhanced in the presence of IFN-gamma. Neither mAb5D3 nor LPS were able to activate macrophages from the LPS-hyporesponsive C3H/HeJ mouse, although these cells responded normally to heat-killed Listeria monocytogenes. The results of several experiments establish that the observed LPS-like activity of mAb5D3 was not due to contaminating endotoxin: 1) the activity of mAb5D3 but not LPS was heat labile at 100 degrees C; 2) the activity of LPS but not mAb5D3, was inhibited by addition of polymyxin B; and 3) quantitative estimates of endotoxin contamination by Limulus amoebocyte lysate reactivity. These experiments thus demonstrate that mAb5D3 can serve as an agonist for LPS-dependent macrophage responses and, when considered with those of our companion paper showing specificity of mAb5D3 for the 80-kDa LPS-binding protein, provide strong support for the concept that the 80-kDa LPS-binding protein previously identified serves as a functional receptor for LPS on murine macrophages.