Non-treated PC-3 and PDGF (HZ-1215) treated PC-3 cells were subjected to SDS PAGE followed by western blot with 29000-1-AP (Phospho-TSC2 (Thr1462) antibody) at dilution of 1:1000 incubated at 37℃ for 1 hour.

Non-treated NIH/3T3 and PDGF (HZ-1215) treated NIH/3T3 cells were subjected to SDS PAGE followed by western blot with 29000-1-AP (Phospho-TSC2 (Thr1462) antibody) at dilution of 1:1000 incubated at 37℃ for 1 hour.

Non-treated and Calyculin A treated cell lysates were subjected to SDS PAGE followed by western blot with 67558-1-Ig (Phospho-GSK3B (Ser9) antibody) at dilution of 1:5000 incubated at room temperature for 1.5 hours. The membrane was stripped and reblotted with HRP-conjugated GAPDH Monoclonal antibody (HRP-60004) as loading control.

Non-treated and Calyculin A treated HEK-293 cells were subjected to SDS PAGE followed by western blot with 80596-1-RR (Phospho-mTOR (Ser2448) antibody) at dilution of 1:10000 incubated at room temperature for 1.5 hours. The membrane was stripped and re-blotted with RICTOR antibody (66867-2-Ig) subsequently.

Non-treated and Calyculin A treated HeLa cells were subjected to SDS PAGE followed by western blot with 80596-1-RR (Phospho-mTOR (Ser2448) antibody) at dilution of 1:10000 incubated at room temperature for 1.5 hours. The membrane was stripped and re-blotted with RICTOR antibody (66867-2-Ig) subsequently.

Non-treated and Calyculin A treated HEK-293 cells were subjected to SDS PAGE followed by western blot with 80698-1-RR (Phospho-TSC2 (Thr1462) antibody) at dilution of 1:5000 incubated at room temperature for 1.5 hours. The membrane was stripped and re-blotted with mTOR antibody (66888-1-Ig) as the loading control.

Non-treated and Calyculin A treated HEK-293 cells were subjected to SDS PAGE followed by western blot with 81654-1-RR (Phospho-TSC2 (Ser939) antibody) at dilution of 1:2500 incubated at room temperature for 1.5 hours. The membrane was stripped and re-blotted with mTOR antibody (66888-1-Ig) as the loading control.

Non-treated MCF-7 cells and Cobalt Chloride treated MCF-7 cells were subjected to SDS PAGE followed by western blot with 28757-1-AP (Phospho-FOXO1 (Ser319) antibody) at dilution of 1:500 incubated at room temperature for 1.5 hours. The membrane was stripped and re-blotted with Beta Actin antibody as loading control.

Immunohistochemical analysis of paraffin-embedded human liver cancer tissue slide using 67558-1-Ig (GSK3B-phospho-S9 antibody) at dilution of 1:300 (under 40x lens). Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0).

Immunofluorescent analysis of (-20°C Ethanol) fixed Calyculin A treated PC-3 cells and non-treated PC-3 cells using Phospho-GSK3B (Ser9) antibody (67558-1-Ig, Clone: 1C9E2 ) at dilution of 1:200 and CoraLite®488-Conjugated AffiniPure Goat Anti-Mouse IgG(H+L).

Immunofluorescent analysis of (4% PFA) fixed PMA treated HEK-293 cells using Phospho-mTOR (Ser2448) antibody (80596-1-RR, Clone: 3L18 ) at dilution of 1:1000 and CoraLite®488-Conjugated AffiniPure Goat Anti-Rabbit IgG(H+L), CL594-phalloidin (red).

TSC2 fusion protein Ag19402 SDS-PAGE