Antibody for hif 1α

Cells in culture or xenograft tissues were lysed with 1% Triton X-100-containing RIPA buffer supplemented with protease/phosphatase inhibitor cocktail from Roche Diagnostics (Indianapolis, IN, USA). Protein concentrations were quantified with the Pierce BCA Protein Assay Kit (Thermo Fisher Scientific). For analysis of GLUT1 (a membrane-associated hydrophobic protein), protein samples were solubilized in sodium dodecyl sulfate (SDS) Laemmli buffer and incubated at 65 °C for 10 min instead of boiling to prevent heat-induced aggregation of hydrophobic proteins. Equal amounts of proteins were resolved by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The antibodies against GLUT1 (catalog #12939), phospho-AMPKα (catalog # 2535), and β-actin (catalog # 4970) were purchased from Cell Signaling Technology (Danvers, MA, USA). The antibody for HIF-1α (catalog # 610958) was from BD Biosciences (San Jose, CA, USA). Immunocomplexes were visualized by an enhanced chemiluminescence detection kit (Thermo Fisher Scientific) using horseradish peroxidase-conjugated secondary antibodies (Cell Signaling Technology).

Matrine was purchased from Sigma-Aldrich (Catalog No. M5319, purity above 97%, Saint Louis, MO, USA). Antibody for HIF-1α was purchased from BD Bioscience (Catalog No. 610958, San Diego, CA, USA). 2-(N-(7-Nitrobenz-2-oxa-1,3-diazol-4-yl)Amino)-2-deoxyglucose (2-NBDG) was purchased from Thermos Fisher Scientific (Eugene, OR, USA). All other reagents were from Sigma-Aldrich (Shanghai, China) unless stated otherwise.