Whole cell lysates were extracted into RIPA buffer with protease inhibitor (Roche). Lysates were spun at max RPM for 45 min at 4°C. Supernatant was collected and quantified for normalization using Pierce BCA protein assay kit (Thermo Fisher Scientific). Than 4x Laemmli sample buffer (Bio-Rad) with 2-mercaptoethanol was added. Samples were not boiled and 20 μg lysate per well was used. Western blotting of these lysates was performed using rabbit polyclonal anti-SLC26A3 ab83545 (Abcam, Cambridge, MA, USA) and rabbit polyclonal anti-β-actin (Abcam, Cambridge, UK).
To localize and quantify SLC26A3 expression paraffin fixed human and mouse colon tissue was processed for microscopy as described18 (link). Both human and mouse samples were stained with rabbit polyclonal anti-SLC26A3 ab244452 (Abcam) followed by Alexa Fluor 488 anti-rabbit secondary Ab (Invitrogen) and counter stained with ProLong Gold Antifade with DAPI (Thermo Scientific).
To localize and quantify SLC26A3 expression paraffin fixed human and mouse colon tissue was processed for microscopy as described18 (link). Both human and mouse samples were stained with rabbit polyclonal anti-SLC26A3 ab244452 (Abcam) followed by Alexa Fluor 488 anti-rabbit secondary Ab (Invitrogen) and counter stained with ProLong Gold Antifade with DAPI (Thermo Scientific).