Anti phospho erk1 2 thr202 tyr204 clone e10

Cells were fixed with 2% formaldehyde (Applichem Lifescience), permabilized with 100% methanol (VWR BDH Prolabo), and stained with anti-Phospho-Akt (Thr308) clone C31E5E and anti-Phospho-Erk1/2 (Thr202/Tyr204) clone E10 from Cell Signaling Technologies following manufacturers' recommendations. RPE-coupled goat anti-mouse and anti-rabbit secondary antibodies (Life Technologies) were used and samples were acquired in a FACSVerse cytometer (BD). FCS files were analyzed in FlowJo software (TriStar).

The following mouse monoclonal antibodies were used as primary antibodies: anti-phospho-ERK1/2 (Thr202/Tyr204) (clone E10, #9106; Cell Signaling Technologies, MA, USA), anti-β-actin (clone AC-15, #A5441; Sigma-Aldrich Corporation, MO, USA), anti-desmoplakin (clone 11-5F; gift from David Garrod, University of Manchester, UK), anti-keratin 14 (clone LL001; Purkis et al., 1990 (link)), anti-keratin 10 (clone DE-K10, #M7002; Agilent Technologies, CA, USA), anti-E-cadherin (clone 36, #610182; BD Biosciences, CA, USA) and anti-Ki-67 (clone MM1, #NCL-L-Ki67-MM1; Leica Biosystems, IL, USA). The following rabbit monoclonal or polyclonal antibodies were also used as primary antibodies: anti-phospho-AKT (Ser473) (clone D9E, #4060), anti-AKT (clone C67E7, #4691), anti-phospho-ERK1/2 (Thr202/Tyr204) (clone D13.14.4E, #4370), anti-ERK1/2 (clone 137F5, #4695) and anti-YAP (clone D8H1X, #14074) were purchased from Cell Signaling Technologies (MA, USA). Secondary antibodies used were as follows: goat anti-rabbit IgG (H+L) Alexa Fluor 594 (#A-11037) and goat anti-mouse IgG (H+L) Alexa Fluor 568 (#A-11031) were purchased from Thermo Fisher Scientific (MA, USA), anti-mouse IgG (H+L), HRP conjugate (#W4021) and anti-rabbit IgG (H+L), HRP conjugate (#W4011) were purchased from Promega Corporation (WI, USA).