Western Blot Analysis of Signaling Proteins

Total cell lysate (25 µg protein) was resolved by SDS-PAGE analysis [30 (link)] using 12% acrylamide gels. Antibodies for western blotting were anti-: c-Raf pS259 (polyclonal), c-Raf pS289/296/301 (polyclonal), VAV1 (polyclonal), Lyn (clone C13F9), SFK pY416 (polyclonal), LCN2 (clone D4M8L), MMP9 (clone D6O3H), Smad2 pS245/250/255 (polyclonal), GAPDH (clone D16H11), HRP anti-mouse, and HRP anti-rabbit (Cell Signaling, MA, USA), c-Raf pS621 (polyclonal) (Thermo Fisher), c-Raf (clone 53) and CD38 (clone 22) (BD Pharmingen, San Jose, CA, USA), c-Cbl pY674 (clone EPR2227) (Abcam, MA, USA) and c-Cbl (polyclonal) (Santa Cruz, CA, USA). All antibodies were diluted 1:1000.

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Bunaciu R.P., MacDonald R.J., Gao F., Johnson L.M., Varner J.D., Wang X., Nataraj S., Guzman M.L, & Yen A. (2017). Potential for subsets of wt-NPM1 primary AML blasts to respond to retinoic acid treatment. Oncotarget, 9(3), 4134-4149.

Publication 2017

c-Raf pS259 c-Cbl

Acrylamide Antibodies C raf Cell Clone Gapdh Gels Mmp9 Mouse Protein Rabbit Sds page Smad2 Vav1

Corresponding Organization :

Other organizations : Cornell University, City University of Hong Kong

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Variable analysis

independent variables

None explicitly mentioned

dependent variables

Protein expression levels of c-Raf pS259, c-Raf pS289/296/301, VAV1, Lyn, SFK pY416, LCN2, MMP9, Smad2 pS245/250/255, c-Raf pS621, c-Raf, CD38, c-Cbl pY674, c-Cbl

control variables

Protein amount (25 µg) for total cell lysate
SDS-PAGE analysis using 12% acrylamide gels
GAPDH as a loading control

positive controls

None explicitly mentioned

negative controls

None explicitly mentioned

Annotations

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