Anti erk1 2 polyclonal antibody

Western blot analysis was performed as previously reported48 (link). Briefly, the cells were lysed with Cell Lysis Buffer (Cell Signaling Technology). The proteins were then separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and electrophoretically transferred to Immobilon-P membranes (Millipore, Billerica, MA, USA). The membranes were probed with anti-Drp1 monoclonal antibodies (1:1000; Cell Signaling Technology), anti-Fis1 polyclonal antibodies (1:200; Santa Cruz Biotechnology, Santa Cruz, CA, USA), anti-Mfn1 polyclonal antibodies (1:1000; Cell Signaling Technology), anti-Mfn2 monoclonal antibodies (1:1000; Cell Signaling Technology), anti-Opa1 monoclonal antibodies (1:1000; BD Biosciences), anti-ERK1/2 polyclonal antibodies (1:1000; Cell Signaling Technology), anti-phospho ERK1/2 (Thr202/Tyr204) monoclonal antibodies (1:2000; BD Biosciences), and anti-β-actin monoclonal antibodies (1:5000; Sigma-Aldrich). The membranes were then incubated with secondary antibodies against rabbit or mouse IgG conjugated to horseradish peroxidase (Cell Signaling Technology). The bands were visualized using an ECL Western Blotting Analysis System (GE Healthcare, Buckinghamshire, UK). Images were acquired using an LAS-3000 Imager (FUJIFILM, Tokyo, Japan).

Western blot analysis was performed as previously reported^{52 (link)}. Briefly, the cells were lysed with cell lysis buffer (Cell Signaling Technology, Danvers, MA, USA), the proteins were separated using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and then electrophoretically transferred to Immobilon-P membranes (Millipore, Billerica, MA, USA). Then, the membranes were probed with anti-ERK1/2 polyclonal antibodies (1:1000, Cell Signaling Technology), anti-phospho ERK1/2 (Thr202/Tyr204) monoclonal antibodies (1:2000, BD Biosciences), anti-Mfn1 monoclonal antibodies (1:1000, Cell Signaling Technology), and anti-β-actin monoclonal antibodies (1:5000, Sigma-Aldrich). The membranes were then incubated with secondary antibodies against rabbit or mouse horseradish peroxidase-conjugated immunoglobulin G (IgG, Cell Signaling Technology). The bands were visualized using an enhanced chemiluminescence (ECL) western blotting analysis system (GE Healthcare, Buckinghamshire, UK) and the images were acquired using an LAS-3000 imager (FUJIFILM UK Ltd., Systems, Bedford, UK).

Protein extraction and quantification were performed as previously described37 (link). Primary antibodies were as follows: anti-p21 monoclonal antibody (Clone SX118, Dako, Denmark), anti-p53 monoclonal antibody (Clone DO-7, Dako), anti-cyclin D1 monoclonal antibody (clone DCS-6, Novocastra, Wetzlar, Germany), and anti-β-actin monoclonal antibody (Clone AC-40, Sigma, Saint Louis, MO, USA), anti-p27 polyclonal antibody (Becton-Dickinson), anti-p16 polyclonal antibody (Santa Cruz Biotechnology, Santa Cruz, CA, USA), anti-ERK1/2 polyclonal antibody (Cell Signaling Technology, Beverly, MA), anti-kRas polyclonal antibody (Santa Cruz Biotecnology), anti-pR polyclonal antibody (Cell Signaling Technology), anti phospo-Rb^{(Ser 780)} polyclonal antibody (Cell Signaling Technology), anti-p38 polyclonal antibody (Abcam, San Francisco, CA), anti-Akt polyclonal antibody (Cell Signaling Technology). Immunoreactivities were revealed with the EnVision dextran polymer visualization system (Dako). Membrane were washed and incubated with ECL (Amersham). Signal acquisition was done with Chemidoc scanner, (BioRad, Hercules, CA) and signals were quantified using a specific densitometric software (Image Lab, BioRad) in absorbance units after light calibration with a reference autoradiography.