Cbr1 levels and Doxol-forming activity in ammonium sulfate-fractionated mouse liver cytosol. (A) Coomassie-stained SDS-PAGE analysis of proteins in ammonium sulfate fractions; 10 µg of protein loaded per lane. P, pellet; S, supernatant. (B) Immunoblot analysis of Cbr1 in ammonium sulfate fractions; 10 µg of protein loaded per lane. (C) Relative amount of Doxol produced by each fraction. Reaction conditions were 200 µM NADPH, 200 µM Dox, and 12 µg fraction protein; incubations were for 1 hour at 37°C. Doxol levels were determined by LC-MS/MS. The relative level of Doxol generated is reported in arbitrary units (AU), where the average level in whole cytosol is set to 2200. Error bars represent 1 S.D.; n = 3 for all reactions; na, not assayed; sup, supernatant.