The deposition of iC3b on injured BM stroma requires IgM. Macrophage-depleted BM stroma cultures were irradiated with 1750 cGy from a cesium-137 source. The gray regions in all panels represent irradiated BM stroma in the absence of serum that were stained with the respective secondary Abs as controls for specificity. (A) Irradiated BM stromal cells were incubated with fresh C57BL/6 NMS (solid line) or heat-inactivated NMS (dotted line) and stained with affinity-purified FITC-conjugated anti-C3 Ab. (B) Fresh Rag-2^-/- serum (dotted line), purified IgM (dashed line), or heat-inactivated NMS (solid line) were incubated with irradiated stromal cells and then incubated with Rag-2^-/- serum as a C source. The cells were then stained with anti-C3Ab to detect iC3b deposition. (C) Following incubation with NMS, irradiated BM stromal cells were stained with goat anti-mouse IgM (solid line) or anti-mouse IgG (not shown). The data are representative of 1 of 3 separate experiments.