F0000000184
ID F0000000184
Title Crude membrane fr. by PM Ext kit
Subcellular Membrane
Protein -
Peptide -
Note For proteomic analysis, cytosol, crude membrane and plasma membrane fractions were prepared from 50 mg of minced liver and kidney using the Plasma Membrane Protein Extraction Kit. The fractionation was performed according to the manufacturer's protocol with slight modifications, i.e., purification by centrifugation at 700 g after the homogenization step was conducted twice to increase the purity of the fractions. For Cyp2b enzyme activity determination, microsomal fraction was prepared from 250 mg of minced liver. The minced liver was homogenized in 1 mL 1.15% KCl solution using a Micro Smash MS-100R (Tomy Seiko, Tokyo, Japan) under cooling. The obtained suspension was centrifuged at 9,000 g for 20 min at 4°C and the supernatant was centrifuged at 105,000 g for 60 min at 4°C. The resulting precipitate was resuspended in suspension buffer (0.1 mM EDTA, 0.25 M sucrose, 10 mM Tris-HCl, pH 7.4).