Fig. 4. Polarised distribution of the GPI-anchored MDP constructs in MDCK cells. (A,B) MDCK cells stably expressing either wtMDP (A) or MDPgly (B) were seeded on polycarbonate filters at confluent levels and left for 3 days to polarise with feeding every other day. Cells were fixed with methanol for 5 minutes prior to labelling with (i) anti-MDP antibody and (ii) anti-Na⁺/K⁺ ATPase antibody; (v) anti-MDP antibody; and (vi) apical marker gp135 antibody; which were visualised using Alexa488- and Alexa594-conjugated secondary antibodies (green and red, respectively). (iii) and (vii) Merged images with colocalisation of antibodies appearing yellow. (iv) and (viii) Z-axis images of cells in (iii) and (vii) are also shown. Bar, 10 µm. The images are representative of five independent experiments.