Catalog no a5441

U87MG and GL261 cells were harvested and lysed in low salt lysis buffer (50 mM Tris-HCl, 150 mM NaCl, 1 mM EDTA, 0.5% NP-40, pH 7.4) supplemented with protease and phosphatase inhibitors (Complete mini EDTA-free and PhosSTOP, Roche Applied Science). Normalized samples were loaded onto 4–12% precast Tris-glycine gradient gels (Life Technologies), subjected to SDS-PAGE, and transferred onto 0.2 μm nitrocellulose membranes (BioRad). Immunoblotting was performed using monoclonal antibodies against transferrin receptor (ThermoFisher, Catalog No. 13-6800) and actin (Sigma, Catalog No. A5441) each at 1:1000 dilution. Protein bands were visualized following incubation with Li-Cor fluorescent secondary antibodies (Li-Cor, Catalog No. P/N 925-32212 and P/N 925-68072) at 1:20,000 dilution using a Li-Cor Odyssey Infrared Imaging System (Li-Cor) and quantified using Odyssey software (Li-Cor). Actin was used as a loading control. Protein band intensity quantified using Li-Cor Odyssey software. Intensity of bands averaged over three separate experiments. Uncropped blots are included in Supplementary Information Section.