FIG 3.

Alloactivation enhances LFA-1-mediated adhesion to MHC-mismatched fibroblasts and interaction of ADAP with LFA-1 and SLP-76. (A) Characterization of in vitro-generated and expanded fibroblasts by flow cytometry with an antibody specific for mouse embryonic fibroblasts (mEF) in comparison to HeLa cells and NIH 3T3 fibroblasts. (B) Syngeneically (syn) or allogeneically (allo) stimulated T cells were seeded on plates cultured with syngeneic eGFP-expressing fibroblasts. The mixed cultures were incubated with anti-human IgG (negative control) or anti-mouse LFA-1 antibody (each at 20 μg/ml) for 2 h at 37°C. After washing and fixing, the adherent T cells were probed with antibody against CD8 and, together with fibroblasts, were visualized by fluorescence microscopy. (C and D) Lysates of syngeneically (syn) or allogeneically (allo) stimulated T cells were immunoprecipitated (IP) and probed (WB) with antibodies as indicated.