Anti aqp2

Kidney sections and fixed cells were immunostained as described previously [10 (link)]. For immunochemical staining, sections were stained with anti-AQP2 (Cat. No. AQP-002; Alomone Laboratories) antibody. Hematoxylin was used for counter-staining. For immunofluorescence, the following antibodies were used: anti–acetylated (ac)-α-tubulin (Cat. No. T7451; Sigma-Aldrich), anti–Na/K-ATPase (Cat. No. ab76020; Abcam), anti-AQP1 (Cat. No. AQP-001; Alomone Laboratories), and anti-AQP2 (Cat. No. AQP-002). To detect cell nuclei, DAPI was applied to samples. Images were captured using a Leica microscope (DM2500; Wetzlar).

Kidney sections were deparaffinized and rehydrated, and then washed with phosphate-buffered saline (PBS) for 5 min each. The sections were incubated in PBS containing 0.1% sodium dodecyl sulfate (SDS; Sigma) for 1 min and washed in PBS for 10 min. To expose the antigen epitope, the sections were boiled in 10 mM sodium citrate buffer (pH 6.0) for 10 min, cooled for 20 min, and then washed three times with PBS for 5 min. The sections were blocked with 1% bovine serum albumin in PBS (blocking buffer) for 30 min and then incubated with anti-ac-α-tubulin, anti-AQP-1 (Alomone Labs, Jerusalem, Israel), and anti-AQP-2 (Alomone Labs) antibodies diluted in blocking buffer overnight at 4 °C. After washing, the sections were incubated with FITC-conjugated goat anti-mouse IgG (Vector Laboratories, Burlingame, CA, USA) or goat anti-rabbit IgG (Vector Laboratories) for 60 min, and then washed three times with PBS for 5 min. To stain cell nuclei, 4′-6-diamidino-2-phenylindole (DAPI; Sigma) was placed on the sections for 1 min.
To detect fragments of primary cilia in the urines, slide glasses were smeared urines, fixed, immunostained using anti-ac-α-tubulin and -Arl13b antibodies, and then observed under a Leica microscope.