3h digoxin

Enisamium iodide and N-methyl-4-N-ethanolpyridinium iodide (internal standard for LC-MS/MS) were provided by Farmak JSC.
Dulbecco’s Modified Eagle’s Medium (DMEM) was obtained from Pan Biotech (Aidenbach, Germany). [³H]-propranolol, rhodamine 123, Krebs-Ringer buffer (KRB), cyclosporine A and [³H]-digoxin from Sigma-Aldrich. Trypsin/ethylenediaminetetraacetic acid (EDTA) solution and sodium dodecyl sulfate from Merck (Darmstadt, Germany). Fluorescein was purchased from VWR International (Darmstadt, Germany). Culture flasks were obtained from VWR International and Transwell filter inserts from Corning Life Sciences (Lowell, MA, USA).

Hank’s balanced salt solution (HBSS), 2-(N-morpholino) ethanesulfonic acid hydrate (MES), N-(2-hydroxyethyl)piperazine-N’-(2-ethanesulfonic acid) (HEPES), 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) (MTT), fluorescein isothiocyanate-conjugated dextran 4 kDa (FD-4), verapamil, [³H]-digoxin, mucin from porcine stomach type III, ML-7 (an MLCK inhibitor), RO-318220 (a PKC inhibitor), genistein (a tyrosine kinase inhibitor), Na⁺/K⁺ ATPase and 4’,6-diamidino-2-phenylindole dihydrochloride (DAPI) were purchased from Sigma-Aldrich (St. Louis, MO, USA). The clear-sol I scintillation cocktail and protease inhibitor cocktail were from Nacalai Tesque (Kyoto, Japan). The P-gp-Glo™ assay and P-gp antibody were obtained from Promega (Madison, WI, USA) and Alexis Biochemicals (San Diego, CA, USA), respectively. Primary antibodies against zonula occludens-1 (ZO-1) and occludin were from Invitrogen (San Diego, CA, USA). Goat anti-mouse IgG H&L Alexa Fluor® 488 and 568 secondary antibodies, FITC-labeled anti-P-gp [UIC2] monoclonal antibody and horseradish peroxidase (HRP)-conjugated secondary antibody were obtained from Abcam (Cambridge, UK). All other chemical reagents were analytical grades.