Immunologic identification of D dimer-induced DNA binding proteins. Subconfluent rat lung fibroblasts were incubated in 0.4% serum for 48 hours followed by exposure to D dimer (1 μmol/L for 45 minutes) or serum-free media controls (SFM). Nuclear proteins were extracted and analyzed by EMSA with 32P-PAIcon (A and B; PAIcon) or 32P-PAIwt (C) as indicated below the autoradiograms. The effect of antibodies that are broadly cross-reactive to fos/jun proteins and to specific fos/jun family members on the protein-DNA binding was assessed through an examination of the competition or supershifting of complex B in the EMSA autoradiogram. (A) Increasing titer of fos (lanes 3 through 6) and jun (lanes 7 through 10) family antibodies from 1 to 2,000 ng/binding reaction. Antibody to CD11b serves as an irrelevant control antibody. (B) Left autoradiogram, indicated antibodies to specific fos family members, with negative control (CREB-2 Ab). Right autoradiogram, indicated antibodies to specific fos family members, with negative control (ATF-2 Ab). (C) Indicated antibodies to specific fos and jun family members, with 32P-labeled PAIwt.
