Fig. 6. Double-staining for CEACAM1 and occludin. Confluent, polarized CEACAM1-L- or CEACAM1-S-expressing MDCK cells were treated with or without hyperosmotic sucrose, fixed with 3% paraformaldehyde and permeabilized by methanol for 2 minutes at room temperature. They were then double-stained with an anti-occludin antibody and Alexa488-conjugated secondary antibodies (green colour) and anti-CEACAM1 (CC16) and Alexa546-conjugated secondary antibodies (red colour). The z-reconstructions show the stainings for both CEACAM1 (red) and occludin (green). The xy-fields show focal planes at the level of the tight junctions; only the staining for occludin (green) is shown in these fields. The tight junctions were intact after treatment with hyperosmotic sucrose, as judged by the occludin staining. Note also that the lateral staining for CEACAM1-L is weaker after sucrose treatment. Bars: 20 µm.

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