Western Blot Analysis of Protein Expression

Western blots were performed as previously described (17 (link), 18 (link), 58 (link)) using NuPage 4 to 12% bis-tris gels and the following antibodies (at the manufacturer’s recommended concentrations): anti-TRPM7 (mouse, clone S74-25, Thermo Fisher Scientific MA5-27620), anti-Cdc42 (rabbit, clone 11A11, Cell Signaling Technology 2466), and anti-IQGAP1 (mouse, clone 9, Santa Cruz Biotechnology, sc-376021) with glyceraldehyde-3-phosphate dehydrogenase (rabbit, clone 14C10, Cell Signaling Technology 2118) or β-actin (mouse, clone C4, BD Biosciences 612656) as a loading control. Secondary antibodies (used at 1:1000 dilution): anti-mouse immunoglobulin G (IgG) horseradish peroxidase (HRP)–linked antibody (Cell Signaling Technologies) and anti-rabbit IgG HRP-linked antibody (Cell Signaling Technologies).

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Yankaskas C.L., Bera K., Stoletov K., Serra S.A., Carrillo-Garcia J., Tuntithavornwat S., Mistriotis P., Lewis J.D., Valverde M.A, & Konstantopoulos K. (2021). The fluid shear stress sensor TRPM7 regulates tumor cell intravasation. Science Advances, 7(28), eabh3457.

Publication 2021

anti-Cdc42 anti-IQGAP1 glyceraldehyde-3-phosphate dehydrogenase β-actin anti-mouse immunoglobulin G (IgG) horseradish peroxidase (HRP)–linked antibody anti-rabbit IgG HRP-linked antibody

Actin Anti antibody Antibodies Antibody Bis tris Cdc42 Clone Dilution Gels Glyceraldehyde 3 phosphate dehydrogenase Horseradish peroxidase Immunoglobulin g Mouse Rabbit Trpm7 Western blots

Corresponding Organization :

Other organizations : Johns Hopkins University, University of Alberta, Pompeu Fabra University, Auburn University, Johns Hopkins Medicine

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Variable analysis

independent variables

None specified

dependent variables

Protein expression levels of TRPM7, Cdc42, and IQGAP1

control variables

Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) or β-actin as loading controls

controls

Positive control: None specified
Negative control: None specified

Annotations

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