Rabbit anti na k atpase antibody

1,2-Dioctanoyl-sn-glycerol (DOG) and 2-dihexanoyl-sn-glycero-3-phospho-l-serine (DPS) were obtained from Avanti Polar Lipids (Alabaster, AL). Phorbol 12-myristate 13-acetate (PMA) was purchased from LC Laboratories (Woburn, MA). Phorbol-12,13-dibutyrate (PDBu) was obtained from Sigma-Aldrich (St. Louis, MO). Deuterated dodecylphosphocholine (d₃₈-DPC), DMSO (d₆-DPC), and ¹⁵NH₄Cl were obtained from Cambridge Isotope Laboratories, Inc. (Tewksbury, MA). Protein quantification was conducted using the Bradford protein assay from Bio-Rad (Hercules, CA) and the BCA (bicinchoninic acid) protein assay kit from Thermo Fisher Scientific (Grand Island, NY). Glutathione sepharose 4B was obtained from GE Healthcare Life Sciences (Piscataway, NJ). HT22 cells were purchased from ATCC (Manassas, VA). Fetal bovine serum (FBS) was from ZenBio (Research Triangle Park, NC). Rattus norvegicus Munc13-1 conjugated with green fluorescent protein (GFP) was a generous gift from N. Brose (Max Planck Institute for Experimental Medicine, Gottingen, Germany). The rabbit anti-GFP antibody, rabbit anti-Na/K-ATPase antibody, rabbit anti-β-actin, and HRP (horseradish peroxidase)-conjugated rabbit anti-IgG antibody used for the Western blot analysis were obtained from Cell Signaling (Danvers, MA). All other reagents were purchased from either MilliporeSigma (Burlington, MA) or Thermo Fisher Scientific.

Cells were transfected for 48 hours, and lysed in lysis buffer (50 mM Tris-HCl, pH7.5, 150 mM NaCl, 2 mM EDTA, 1% Nonidet P-40, and protease inhibitor cocktail) (Roche, USA)) to prepare cell extracts for Western blot analysis and Co-IP analysis as described previously [17 (link), 36 (link), 37 ]. A rabbit anti-Na_v1.5 antibody (Alomone Labs, Jerusalem BioPark) was used at a dilution factor of 1:1000. A mouse anti-tubulin (Millipore, USA) antibody was used at a dilution factor of 1:5000. A goat anti-rabbit HRP-conjugated secondary antibody and a goat anti-mouse HRP-conjugated secondary antibody were all from Millipore (USA) and used at a dilution factor of 1:20,000. A rabbit anti-UBC9 antibody (Santa Cruz, USA) was used at a dilution factor of 1:1000. A mouse anti-Myc antibody (MBL, JAPAN) was used at a dilution factor of 1:1000. A rabbit anti-Na,K-ATPase antibody (Cell Signaling Technology, USA) was used at a dilution factor of 1:1000. A mouse anti-FLAG antibody, a rabbit anti-HA antibody and a rabbit anti-GFP antibody were all from MBL (JAPAN). The goat anti-rabbit IgG and goat anti-mouse IgG were from Santa Cruz Biotechnology. The anti-ubiquitin mouse monoclonal antibody FK2 (BMLPW8810, Enzo Life Science) was used at a dilution factor of 1:800.