Fig. 4.

Fluorescence microscopic analysis of T84 monolayers treated with IFN-γ. Cultures were stained with antibodies against E-cadherin (red) and TG2 (green) proteins, as well as with a streptavidin conjugate (blue) that recognizes 5BP incorporated at any site where extracellular TG2 is activated. Images were generated by using a Carl Zeiss 510 Meta confocal microscope with an oil immersion compatible 40× objective capable of digital imaging correlation. A, treatment of T84 monolayers with 0 to 1000 U/ml IFN-γ for 48 h. B, T84 monolayers exposed to 0 to 1000 U/ml IFN-γ for 48 h were pretreated with 25 μM ERW1041E before addition of 5BP. C, single plane images were generated at different depths of a T84 monolayer treated with 1000 U/ml IFN-γ for 48 h. The depth of each focal plane is illustrated in the upper left corner (0–10 μm) of each single plane image. Images are arranged from apical to basolateral side of the T84 monolayer as read left to right. Each transwell was performed in triplicate (n = 3). All images were processed in the same manner.