Anti prsk antibody

Manufactured by Cell Signaling Technology

Sourced in United States

The Anti-pRSK antibody is a laboratory reagent used to detect the phosphorylated form of the Ribosomal S6 Kinase (RSK) protein. RSK is a serine/threonine protein kinase that plays a role in the Ras/MAPK signaling pathway. The antibody specifically recognizes the phosphorylated form of RSK, which can provide information about the activation state of this signaling pathway in cells or tissues.

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2 protocols using anti prsk antibody

In Vitro Kinase Activity Assay

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A375 or HCT116 cells were seeded at 1.5 × 10⁴ cells per well into 96-well plates and allowed to recover for 16 h, prior to the addition of compounds (in 0.1% DMSO v/v) and incubation for a further 2 or 7 h. Cells were lysed by adding cell lysis buffer (Cell Signaling Technology, Massachusetts, USA) and incubating at room temperature for 20 minutes. Custom MSD plates (Meso Scale Discovery, Maryland, USA) pre-coated with anti-pRSK antibody (Cell Signaling Technology, Massachusetts, USA) were blocked with kit blocking buffer for 1 hour at room temperature, prior to washing. Equivalent amounts of protein lysate were added to the blocked plates and incubated for 3 hours at room temperature. After washing, plates were incubated for 1 hour at room temperature with sulfo-tag conjugated anti-RSK detection antibodies (R&D Systems, Minneapolis, USA). Plates were washed, and read buffer added before reading the plate on a MESO QuickPlex SQ 120 (Meso Scale Discovery, Maryland, USA). IC₅₀ values were calculated using a sigmoidal dose–response equation (Prism GraphPad software, La Jolla, CA, USA).

Lebraud H., Surova O., Courtin A., O'Reilly M., Valenzano C.R., Nordlund P, & Heightman T.D. (2018). Quantitation of ERK1/2 inhibitor cellular target occupancies with a reversible slow off-rate probe †Electronic supplementary information (ESI) available. See DOI: 10.1039/c8sc02754d. Chemical Science, 9(45), 8608-8618.

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Antibody-based Protein Analysis Protocol

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Anti-Eed antibody (#09-774, rabbit polyclonal, 1:1,000) was purchased from Millipore. Anti-H3K27me3 antibody (GTX1121184, rabbit polyclonal, 1:500) was purchased from GeneTex. Anti-Actin antibody (I-19, rabbit polyclonal, 1:500) was purchased from Santa Cruz Biotechnology. Anti-Hif1a antibody (NB100-449, rabbit polyclonal, 1:500) was purchased from Novus Biologicals. Anti-Bnip3 antibody (ab10433, mouse monoclonal (ANa40), 2 μg ml⁻¹) was purchased from Abcam. Anti-p-ERK1/2 antibody (#4370, rabbit monoclonal, 1:1,000), anti-p-MEK1 antibody (#9154, rabbit monoclonal, 1:1,000), anti-p-cRaf antibody (#9427, rabbit monoclonal, 1:1,000), anti-p-RSK antibody (#9335, rabbit monoclonal, 1:1,000) and anti-p-p38 antibody (#4511, rabbit monoclonal, 1:1,000), anti-total ERK antibody (#9102, rabbit polyclonal, 1:1,000), anti-p-Smad1/5/8antibody (#9511, rabbit polyclonal; 1:1,000), anti-Smad2 antibody (#5339, rabbit monoclonal, 1:1,000), anti-p-Smad2 antibody (#3104, rabbit polyclonal, 1:1,000), anti-TGF-β receptor II antibody (#3713, rabbit polyclonal, 1:1,000), anti-p-STAT1 antibody (#9171, rabbit polyclonal, 1:1,000) and anti-pSTAT3 antibody (#9145, rabbit monoclonal, 1:2,000) were purchased from Cell Signaling Technology. Western blot analysis was performed according to the standard procedure.

Mirzamohammadi F., Papaioannou G., Inloes J.B., Rankin E.B., Xie H., Schipani E., Orkin S.H, & Kobayashi T. (2016). Polycomb repressive complex 2 regulates skeletal growth by suppressing Wnt and TGF-β signalling. Nature Communications, 7, 12047.

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