Phosphoprotein Analysis of IP6 Treatment

Western blot analysis was performed as described previously [15 (link)]. MG63.3 cells were treated with IP6 (0 or 5 mM) for 6 hours then lysed in SDS gel loading buffer. Lysates from equal number of cells were loaded on 4-20% SDS-PAGE gels. After electrophoresis, proteins were transferred to a nitrocellulose membrane and probed with primary antibodies against phospho-Akt (Ser473), phospho-GSK3α/β (Ser21/9) and phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (Cell Signaling Technology, Inc. Boston, MA). β-Actin (Sigma, St. Louis, MO) was used as a loading control.

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Ren L., Hong E.S., Mendoza A., Issaq S., Hoang C.T., Lizardo M., LeBlanc A, & Khanna C. (2017). Metabolomics uncovers a link between inositol metabolism and osteosarcoma metastasis. Oncotarget, 8(24), 38541-38553.

Publication 2017

phospho-Akt (Ser473), phospho-GSK3α/β (Ser21/9) phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) β-Actin

Actin Antibodies Buffer Cells Electrophoresis Erk1 Gels Gsk3α Membrane Nitrocellulose Proteins Sds page Western blot analysis

Corresponding Organization :

Other organizations : National Cancer Institute, Center for Cancer Research

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Variable analysis

independent variables

IP6 (0 or 5 mM)

dependent variables

Phosphorylation levels of Akt (Ser473), GSK3α/β (Ser21/9), and p44/42 MAPK (Erk1/2) (Thr202/Tyr204)

control variables

Equal number of MG63.3 cells
Loading control: β-Actin

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