<p>MDCK cells stably expressing PrP<sup>C</sup>WT, PrP<sup>C</sup>G1, PrP<sup>C</sup>G2 or PrP<sup>C</sup>G3 were grown in Transwells for 4 to 5 days until they were fully polarized. (A) Cells were separately stained with the 3F4 antibody (green) followed by permeabilisation and staining with an antibody against ZO-1 (red), a constituent of tight junctions, indicating the cell polarity. Confocal microscopy of a Z-stack of PrP<sup>C</sup>WT (left) at the level of tight junctions stained with ZO-1, and YZ-sections (right) of all glycomutants indicate both the integrity of the polarized monolayer and a redistribution of PrP<sup>C</sup>G1 and PrP<sup>C</sup>G2 to the apical compartment when compared to PrP<sup>C</sup>WT and PrP<sup>C</sup>G3. Localization of the apical (a) and basolateral (b) compartment is indicated. (B) After immunocytochemistry under non-permeabilising conditions with the 3F4 antibody, serial Z-stacks from the bottom to the top were taken with confocal microscopy. YZ images shows transversal cut trough cells at the mid level, marked with a dashed line. PrP<sup>C</sup>WT and PrP<sup>C</sup>G3 were mainly found in the basolateral compartment whereas PrP<sup>C</sup>G1 and PrP<sup>C</sup>G2 were mainly found in both compartments. Scale bars represent 10 µm.</p
