Rc dctm kit

Liver samples were homogenized in an ice-cold buffer (7 M urea, 2 M thiourea, 2% CHAPS, 50 mM DTT, 0.8% CA, 1 mM phenylmethylsulfonyl fluoride (PMSF), Bio-Rad, USA). The homogenates were centrifuged at 15,000 × g for 30 min at 4 °C. Total liver protein concentration was determined using an RC-DC^TM kit (Bio-Rad, USA).

Protein extraction from mammary tissues was performed according the method described by Duanmu et al. (13 (link)). The mammary tissue separately samples in BG and HG are mixed in equal amounts and washed three times with ice-cold saline containing 1 mM phenylmethylsulfonyl fluoride (PMSF), and then homogenized in ice-cold lysis buffer (2 M thiourea, 7 M urea, 50 mM dithiothreitol (DTT), 2% (w/v) 3-[(3-cholamidopropyl) dimethylammonio]-1-propanesulfonate, 0.5% (v/v) Bio-Lyte Ampholyte and 1 mM PMSF) by 1:5 (w/v). The homogenates were kept at room temperature for 30 min, followed by centrifugation at 15,000 × g for 30 min at 4°C. The HG samples were treated in the same way. The samples were stored at −80°C until analysis. The protein concentration of the supernatant was determined using an RC DCTM kit (Bio-Rad).