Rabbit p akts473

Cells were lysed with RIPA lysis buffer (150 mM NaCl, 1.0% Triton X-100, 0.5% sodium deoxycholate, 0.1% sodium dodecyl sulphate, 50 mM Tris, pH 8.0) supplemented with 100 μl/mL protease inhibitor. Cell lysate was then processed for immunoblotting and visualised with ECL substrate as described^{12 (link)}. The following antibodies have been used: Primary Antibodies: Cell Signalling Technologies (Danvers, MA): Rabbit-Cyclin D (1/1000), Mouse- Cyclin E (HE12) (1/1000), Mouse- Cyclin A (BF683) (1/2000), Rabbit- Caspase-3 (1/1000), Rabbit Cleaved Caspase-3 (Asp175) (1/1000), Rabbit p-S6S235/236 (1/800), Rabbit Total S6 (1/2000), Mouse IgG1 mTOR (1/1000), Rabbit p-mTORS2448 (1/800), Rabbit E2F1 (1/1000), Rabbit pp-RbS780 (1/1000), Rabbit Rb (1/2000), Rabbit Rictor (1/800), Rabbit Raptor (1/800), Rabbit p-AktS473 (1/1000), Rabbit t-Akt (1/2000), Rabbit p-p70S6K (1/500), Rabbit β-Tubulin (1/2000). Santa Cruz (Dallas, TX): Rabbit-poly MCL1 (1/500). Purified Rabbit N-terminal^{45 (link)} (1/2000). Bethyl Labs (Montgomery, TX): Rabbit USP9x C terminal (1/2000) Sigma Aldrich (St Louis, MO): Mouse β-Tubulin (1/2000), Trevigen (Gaithersberg, USA): Rabbit GAPDH (1/2000). Secondary Antibodies: Life Technologies (Mulgrave, AUS): Rabbit HRP (1/5000), Mouse HRP (1/5000). Millipore (Darmstadt, Germany): Rabbit HRP (1/5000), Mouse HRP (1/5000).