Type 1 insulin-like growth factor receptor levels were analyzed by Western blotting performed as described in detail elsewhere (12 (link)) and using a rabbit anti-IGF-IR antiserum (C-20, Santa Cruz Biotechnology, Dallas, TX, United States) and a horseradish peroxidase-conjugated donkey anti rabbit IgG antibody (GE Healthcare Life Sciences, Pittsburgh, PA, United States). To normalize for loading, the membranes were stripped and re-probed with a monoclonal anti-tyrosine tubulin antibody (Sigma-Aldrich, St. Louis, MO, United States). To analyze ERK1 and 2 phosphorylation in response to IGF-I, tumor cells, cultured overnight in serum-free medium, were stimulated with 100 ng/ml IGF-I (US Biological, Salem, MA, United States) for 10 min, lysed on ice in the presence of phosphatase inhibitors and the cell lysates separated on 8.5% SDS-polyacrylamide gels. The blots were probed, first with an anti-phospho-p44/42 ERK (Thr202/Tyr204) antibody and then with an anti p44/42 ERK antibody (Cell Signaling, Whitby, ON, Canada).
Samani A.A., Nalbantoglu J, & Brodt P. (2020). Glioma Cells With Genetically Engineered IGF-I Receptor Downregulation Can Persist in the Brain in a Dormant State. Frontiers in Oncology, 10, 555945.
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