Immunofluorescent detection of FXIII-A in THP-1 cells. (A) The distribution of fluorescence for Ab-2 in fixed permeabilized nonadherent THP-1 cells was predominantly cytoplasmic in certain cells (top left) but predominantly nuclear in other cells (bottom right). (B) Adherent PMA-differentiated THP-1 cells were treated for 6 days with IL-4 (20 ng/mL). After fixation and permeabilization cells were probed with anti–FXIII-A antibody Ab-2 and with antibodies to Golgi markers GM130 (Bi) and TGN46 (Bii). In the majority of cells (shown in top rows in Bi-ii), Ab-2 fluorescence was distributed throughout the cytosol and nucleus. Ab-2 fluorescence was concentrated in the perinuclear area where it overlaid Golgi cisternae, but the regions of greatest intensity of Golgi and Ab-2 fluorescence did not correspond. In occasional cells (shown in bottom rows in Bi-ii), Ab-2 and Golgi marker fluorescence colocalized in podosome-like membrane extrusions (yellow pixels). In a high proportion of cells in which Ab-2 fluorescence was apparent in the periphery, perinuclear FXIII-A fluorescence was notably reduced. (C) THP-1 cells, grown as in panel B, were treated for 1 hour with Brefeldin A (BFA; 10 μg/mL) before fixation and permeabilization. Immunofluorescence for FXIII-A and GM130 remained within peripheral plasma membrane–associated structures despite dispersion of perinuclear Golgi elements to vesicles.