PA1-26655

antibody from Invitrogen Antibodies

Targeting: RAF1 c-Raf, CRAF, Raf-1

Western blot (Supportive data in Antibodypedia)

Antibody Data

Product number

PA1-26655

Provider

Invitrogen Antibodies

Product name

Anti-Phospho-c-Raf (Ser621) Polyclonal Antibody

Antibody type

Polyclonal

Antigen

Synthetic peptide

Description

PA1-26655 detects c-Raf (phospho S621) from human samples. PA1-26655 is expected to cross react with mouse (100% conserved) and rat (100% conserved) due to sequence homology. PA1-26655 has been successfully used in Western blot procedures. The PA1-26655 immunogen is a synthetic phosphopeptide corresponding to a portion of human c-Raf containing serine 621. The sequence is conserved in mouse and rat.

Reactivity

Human, Mouse, Rat

Host

Rabbit

Isotype

IgG

Vial size

50 µL

Concentration

0.5 mg/ml

Storage

-20° C, Avoid Freeze/Thaw Cycles

References

Submitted references

Nuclear Raf-1 kinase regulates the CXCR5 promoter by associating with NFATc3 to drive retinoic acid-induced leukemic cell differentiation.

Geil WM, Yen A
The FEBS journal 2014 Feb;281(4):1170-80

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The Src-family kinase inhibitor PP2 rescues inducible differentiation events in emergent retinoic acid-resistant myeloblastic leukemia cells.

Jensen HA, Styskal LE, Tasseff R, Bunaciu RP, Congleton J, Varner JD, Yen A
PloS one 2013;8(3):e58621

Western blot

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western blot antibody-peptide competition and mutant analysis using a Phospho-c-Raf pSer621 polyclonal antibody (Product # PA1-26655). Immunoprecipitates of HEK293 cells overexpressing wild-type c-Raf and stimulated with 50 ng/ml EGF for 3 minutes (1-4) or c-Raf mutant S621A (5) were resolved by SDS-PAGE on a 10% Tris-glycine gel and transferred to PVDF. The membrane was blocked with a 5% BSA-TBST buffer overnight at 4ºC, and incubated with c-Raf (pS621) antibody for two hours at room temperature in a 3% BSA-TBST buffer, following prior incubation with: the phosphopeptide immunogen (1), a generic phosphoserine-containing peptide (2), the non-phosphopeptide corresponding to the phosphopeptide immunogen (3), or no peptide (4, 5). The inset shows the relative amount of total c-Raf protein in the wild-type vs. the S621A mutant extracts via c-Raf pan antibody. The data shows that only the phosphopeptide corresponding to c-Raf (pS621) blocks the antibody signal and that the antibody does not recognize the S621A mutant, demonstrating the specificity of the antibody.