Association of FXIII-A with TG2 but not protein disulphide isomerase in podosomes. Adherent PMA-differentiated THP-1 cells were treated for 6 days with IL-4 (20 ng/mL) and, after fixation and permeabilization, were probed with antibodies to FXIII-A, to protein disulphide isomerase (PDI), and to TG2. (A) Cytoplasmic full-length FXIII-A does not correspond in distribution to PDI (top row). PDI is excluded from podosomes that show a strong signal for Ab-2 (bottom row, filled arrowhead) but is present on other membrane processes (open arrowhead). (B) Cells were imaged with the use of a low-power objective to detect FXIII-A (green) and TG2 (red) fluorescence over the full depth of the cell. Left panel shows an overlay of red and green fluorescence, whereas right panel shows a triple overlay with DAPI fluorescence. A high proportion of cells preferentially expressed one or other transglutaminase, but some cells coexpressed FXIII-A and TG2. Both enzymes were present in the cytoplasm, whereas FXIII-A, but not TG2, was also present in the nucleus. Scale bars represent 100 μm. (C) In cells that coexpressed FXIII-A and TG2, both antigens colocalized in podosomes. (D) Immunofluorescence for Ab-2 (green) corresponded with immunofluorescence detected with a sheep polyclonal anti–FXIII-A antibody. Cell margins are apparent in an image with enhanced γ correction (γ, +2.5).