#4432 Store at -20°c a-raf Antibody W, ip endogenous H, M, r 68 kDa Rabbit Background



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#74908


#4432

Store at -20°C

A-Raf Antibody

W, IP

Endogenous

H, M, R

68 kDa

Rabbit**

Background: A-Raf, B-Raf and c-Raf (Raf-1) are the main 

effectors recruited by GTP-bound Ras to activate the MEK-

MAP kinase pathway (1). Activation of c-Raf is the best un-

derstood and involves phosphorylation at multiple activating 

sites including Ser338, Tyr341, Thr491, Ser494, Ser497 and 

Ser499 (2). p21-activated protein kinase (PAK) has been 

shown to phosphorylate c-Raf at Ser338 and the Src family 

phosphorylates Tyr341 to induce c-Raf activity (3,4). Ser338 

of c-Raf corresponds to similar sites in A-Raf (Ser299) 

and B-Raf (Ser445), although this site is constitutively 

phosphorylated in B-Raf (5). Inhibitory 14-3-3 binding sites 

on c-Raf (Ser259 and Ser621) can be phosphorylated by Akt 

and AMPK, respectively (6,7). While A-Raf, B-Raf and c-Raf 

are similar in sequence and function, differential regulation 

has been observed (8). Of particular interest, B-Raf contains 

three consensus Akt phosphorylation sites (Ser364, Ser428 

and Thr439) and lacks a site equivalent to Tyr341 of c-Raf 

(8,9). The B-Raf mutation V600E results in elevated kinase 

activity and is commonly found in malignant melanoma 

(10). Six residues of c-Raf (Ser29, Ser43, Ser289, Ser296, 

Ser301 and Ser642) become hyperphosphorylated in a 

manner consistent with c-Raf inactivation. The hyperphos-

phorylation of these six sites is dependent on downstream 

MEK signaling and renders c-Raf unresponsive to subse-

quent activation events (11).  

Specificity/Sensitivity: A-Raf Antibody detects 

endogenous levels of total A-Raf. This antibody does not 

cross-react with c-Raf or B-Raf.

Source/Purification: Polyclonal antibodies are produced 

by immunizing animals with a synthetic peptide (KLH-cou-

pled) corresponding to residues close to the linker domain 

of human A-Raf. Antibodies are purified by protein A and 

peptide affinity chromatography.

Storage: Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM 

NaCl, 100 µg/ml BSA and 50% glycerol. Store at –20°C.  

Do not aliquot the antibody.

*Species cross-reactivity is determined by western blot.

** Anti-rabbit secondary antibodies must be used to 

detect this antibody.

Recommended Antibody Dilutions: 

Western Blotting 

1:1000 

Immunoprecipitation 1:50



For application specific protocols please see the web 

page for this product at www.cellsignal.com.

Please visit www.cellsignal.com for a complete listing 

of recommended companion products.

Orders

n

877-616-CELL (2355)



orders@cellsignal.com

Support

n

877-678-TECH (8324)



info@cellsignal.com

Web

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www.cellsignal.com



Applications

Species Cross-Reactivity*

Molecular Wt.

Source

© 2010 Cell Signaling T

echnology

, Inc.

Background References:

  (1)  Avruch, J. et al. (1994) Trends Biochem. Sci. 19, 

279–283.

  (2)  Chong, H. et al. (2001) EMBO J. 20, 3716–3727.

  (3)  King, A.J. et al. (1998) Nature 396, 180–183.

  (4)  Fabian, J.R. et al. (1993) Mol. Cell Biol. 13, 

7170–7179.

  (5)  Mason, C.S. et al. (1999) EMBO J. 18, 2137–2148.

  (6)  Zimmermann, S. and Moelling, K. (1999) Science 286, 

1741–1744.

  (7)  Sprenkle, A.B. et al. (1997) FEBS Lett. 403, 254–258.

  (8)  Marais, R. et al. (1997) J. Biol. Chem. 272, 

4378–4383.

  (9)  Guan, K.L. et al. (2000) J. Biol. Chem. 275, 

27354–27359.

 (10)  Davies, H. et al. (2002) Nature 417, 949–954.

 (11)  Dougherty, M.K. et al. (2005) Mol. Cell 17, 215–224.

IMPORTANT: For estern blots, incubate membrane with diluted antibody in 5% w/v BSA, 1X TBS, 0.1% Tween-20 

at 4°C with gentle shaking, overnight.

kDa

140


100

80

C6 



 

60

50



40

A-Raf 

HT2


9

NIH/3T


3

Western blot analysis of extracts from HT-29, NIH-3T3 and 

C6 cell lysates, using A-Raf Antibody.

Entrez-Gene ID #369 

Swiss-Prot Acc. #P10398 

Species Cross-Reactivity Key:

H—human

M—mouse

R—rat

Hm—hamster

Mk—monkey

Mi—mink

C—chicken

Dm—D. melanogaster X—Xenopus

Z—zebrafish

B—bovine

Dg—dog Pg—pig Sc—S. cerevisiae Ce—C. elegans

Hr—Horse

All—all species expected

Species enclosed in parentheses are predicted to react based on 100% homology.



Applications Key:

W—Western

IP—Immunoprecipitation

IHC—Immunohistochemistry

ChIP—Chromatin Immunoprecipitation

IF—Immunofluorescence

F—Flow cytometry

E-P—ELISA-Peptide

rev. 01/22/16



For Research Use Only. Not For Use In Diagnostic Procedures.

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