Coo protein dosage kit

T75-cm² flasks of HEK-293 cells were lysed in 1.0 ml of lysis buffer (50 mM HEPES pH 7.4, 150 mM NaCl, 10 % glycerol, 1 % triton, 1 mM EGTA supplemented with 10 mM N-ethylmaleimide and protease inhibitors). Protein concentration was systematically determined by performing a Bradford assay (Coo protein dosage kit; Interchim, Montluçon, France). Eighty micrograms of proteins were loaded on an SDS-PAGE gel. Protein transfer was done with the dry system transfer iBlot® from Invitrogen (Invitrogen, Basel, Switzerland). Immunoblotting was accomplished by using the SNAP id® system of Millipore (Millipore, Zug, Switzerland). Fluorescent secondary antibodies were used, and detection was realized using the LICOR system® (Lincoln, USA). The intensity of the bands was quantified with the Odyssey software (LICOR).

HEK293 cells or homogenized whole mouse heart were lysed in 1.0 mL of lysis buffer (50 mM Hepes pH 7.4, 150 mM NaCl, 10% glycerol, 1% triton, mM EGTA supplemented with 10 mM N-ethyl maleimide and protease inhibitors (Roche, Basel, Switzerland). Protein concentrations were determined by performing Bradford assays (Coo protein dosage kit; Interchim, Montluçon, France). Forty μg of protein was loaded onto a SDS-PAGE gel. Protein transfer was performed with the dry system transfer i-blot from Invitrogen (Invitrogen, Basel, Switzerland), and immunoblotting using the snap-id system of Millipore (Millipore, Zug, Switzerland). Detection was achieved using the LICOR system^®, and the intensity of the bands was quantified with Odyssey software (LICOR, Lincoln, NE, USA).