Rabbit anti β zo 1

Gallbladders from newborn piglets were excised, placed in ice-cold 30% sucrose solution, quick frozen with liquid nitrogen in optical cutting temperature compound, and stored at −80° C. Frozen tissue was then sectioned using a Cryostat microtome (7 μm sections), fixed in 4% formaldehyde (room temperature, 15 minutes), permeabilized with 0.2% Triton X-100 (Thermo Fisher; room temperature, 20 minutes) and blocked in Super-Block (Thermo Fisher) supplemented with 5% filtered normal goat serum (Jackson Immunologicals; room temperature, 1 hour). Sections were then immunostained with rabbit anti-β-ZO-1 (1:100; Zymed) and mouse anti-CFTR 596 (1:100, CFF/UNC) for 2 hours at 37° C. Subsequently, the sections were incubated in goat anti-mouse Alexa Fluor 488 (1:1000; Molecular Probes) and goat anti-rabbit Alexa Fluor 568 (1:1000; Molecular Probes/Invitrogen), mounted in Vectashield plus DAPI (Vector Labs), and visualized using an Olympus Fluoview FV1000 confocal microscope.