Description
Product Usage Information
Western Blotting: 1:1000
Immunoprecipitation: 1:200
Immunohistochemistry (Paraffin): 1:100 - 1:400
Storage
Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/mL BSA, 50% glycerol, and less than 0.02% sodium azide. Store at -20°C. Do not aliquot the antibody. For a carrier free (BSA and azide free) version of this product see product # 60407 .
Protocol
Available protocols: Western Blotting, Immunoprecipitation, Immunohistochemistry (Paraffin)
Specificity / Sensitivity
B-Raf (E3T5C) Mouse Monoclonal Antibody recognizes endogenous levels of total B-Raf protein.
Species Reactivity: Human, Mouse, Rat, Monkey
Source / Purification
Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Gln356 of human B-Raf protein.
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 understood and involves phosphorylation at multiple activating sites, including Ser338, Tyr341, Thr491, Ser494, Ser497, and Ser499 (2). p21-activated 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 (Ser446), 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 (Ser365, Ser429, and Thr440) and lacks a site equivalent to Tyr341 of c-Raf (8,9). Research studies have shown that 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 hyperphosphorylation of these six sites is dependent on downstream MEK signaling and renders c-Raf unresponsive to subsequent activation events (11).
Alternate Names
94 kDa B-raf protein; B-raf; B-Raf proto-oncogene serine/threonine-protein kinase (p94); B-Raf proto-oncogene, serine/threonine kinase; B-Raf serine/threonine-protein; B-RAF1; BRAF; BRAF1; FLJ95109; MGC126806; MGC138284; murine sarcoma viral (v-raf) oncogene homolog B1; NS7; p94; Proto-oncogene B-Raf; RAFB1; Serine/threonine-protein kinase B-raf; v-raf murine sarcoma viral oncogene homolog B; v-Raf murine sarcoma viral oncogene homolog B1
