Fig. 4. CNF-1 stimulates occludin internalization at an ultrastructural level. Confluent control and CNF-1-treated T84 monolayers were processed for cryo-immunoelectron microscopy (A) by labeling occludin with 15 nm gold particles (pseudo-colored in red) and caveolin-1 with 10 nm gold particles (black). In control cells (a), occludin localized mainly to the apical-most part of the basolateral plasma membrane (arrow). In CNF-1 treated cells, there was a reduction in occludin at the plasma membrane of the TJ domain (b, #). Some labeling was now found to co-localize with caveolin-1 in intracellular caveolae and endosomal-like compartments (c). Bar, 100 nm. Following quantitation of occludin (black bars) and caveolin-1 (gray bars) gold particles at the TJ, statistically significant reductions in both proteins were observed in CNF-treated monolayers relative to controls (B).