Phospho akt ser473 rabbit monoclonal antibody

The tissue samples were incubated with 1 : 100 phospho-AKT (Ser473) rabbit monoclonal antibody (Cell Signaling Tech, MA, USA), or a 1 : 100 dilution phospho-p44/42 MAPK (p-ERK1&2, Thr202/Tyr204) rabbit monoclonal antibody (Cell Signaling Tech, MA, USA) overnight at 4–8°C. After washing, the sections were incubated with the Envision system/HRP (DAKO Cytomation, USA) for 30 min at room temperature. The percentage area of p-ERK1&2 and p-AKT immunoreactivated proportionate to the total area was analyzed by ImageJ 1.38 software (National Institutes of Health, USA).

PBK/TOPK mouse monoclonal antibody was purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA), phospho-p44/42 MAPK (Erk1/2) rabbit monoclonal antibody, phospho-AKT (Ser473) rabbit monoclonal antibody, phospho-PTEN (ser380/Thr382/382) mouse monoclonal antibody, and EpCAM (VU1D9) mouse monoclonal antibody conjugated to Alexa Fluor 594 were purchased from Cell Signaling Technology, Inc. (Beverly, MA, USA). Cytokeratin 19 rabbit monoclonal antibody was purchased from Abcam, Inc. (Cambridge, MA, USA), CD45 rabbit monoclonal antibody was purchased from Biosen, Inc. (Beijing, China), goat anti-mouse IgG conjugated to FITC and TRITC were purchased from Biosen, Inc. (Beijing, China), goat anti-rabbit IgG conjugated to FITC and TRITC were purchased from Biosen Inc. (Beijing, China), HRP-labeled goat anti-mouse IgG(H+L) and goat anti-rabbit IgG(H+L) were purchased from SinoBio, Inc. (SuZhou, China), and 4,6-diamino-2-phenyl indole (DAPI) was purchased from Beyotime, Inc. (Beijing, China), Diaminobenzidine (DAB) substrate was purchased from Dako Denmark A/S (Glostrup, Denmark), packaging vectors including pMD2.0G and psPAX were purchased from Addgene Inc. (Cambridge, MA, USA), polybrene were purchased from EMD Millipore (Billerica, MA, USA), puromycin were purchased from Sigma-Aldrich (St Louis, MO, USA).

The following antibodies were used: p44/42 MAPK (ERK1/2) rabbit monoclonal antibody (Cat# 9102, RRID:AB_330744; Cell Signaling Technology, Beverly, MA, USA), phospho-p44/42 MAPK (ERK1/2; Thr202/Tyr204) rabbit monoclonal antibody (Cat# 9101, RRID:AB_331646; Cell Signaling Technology), Akt (pan) (C67E7) rabbit monoclonal antibody (Cat# 4691, RRID:AB_915783; Cell Signaling Technology), phospho-Akt (Ser473) rabbit monoclonal antibody (Cat# 9271, RRID:AB_329825; Cell Signaling Technology), and anti-phospho-AMPKα (Thr172) rabbit monoclonal antibody (Cell Signaling Technology), anti-AMPKα rabbit monoclonal antibody (Cell Signaling Technology), anti-CRBN mouse polyclonal antibody (Abnova, Taipei, Taiwan), anti-AGEs mouse monoclonal antibody (Transgenic, Inc., Kobe, Japan), anti-GAPDH mouse monoclonal antibody (Wako).