BRAF (mutated V600E) Antibody (Mouse mAb) [M8P1]

Application: Reactivity: Human

Usage Information

Dilution
WB1:1000 IHC1:1000

Application
WB, IHC

Reactivity
Human

Source
Mouse Monoclonal Antibody

Storage Buffer
PBS, pH 7.2+50% Glycerol+0.05% BSA+0.01% NaN3

Storage (from the date of receipt)
-20°C (avoid freeze-thaw cycles), 2 years

Predicted MW Observed MW
84 kDa 85 kDa,36 kDa
^*Why do the predicted and actual molecular weights differ? The following reasons may explain differences between the predicted and actual protein molecular weight. Post-translational modifications(e.g., phosphorylation, glycosylation); Splice variants and isoforms; Relative charge; Multimerization.

Datasheet & SDS

Biological Description

Specificity
BRAF (mutated V600E) Antibody (Mouse mAb) [M8P1] detects endogenous levels of total BRAF protein only when muted at V600E.

Clone
M8P1

Synonym(s)
BRAF1, RAFB1, BRAF, Serine/threonine-protein kinase B-raf, Proto-oncogene B-Raf, p94, v-Raf murine sarcoma viral oncogene homolog B1

Background
BRAF V600E is an oncogenic variant of the RAF family serine/threonine kinase BRAF in which a valine-to-glutamate substitution at codon 600 within the activation segment of the kinase domain produces a constitutively active enzyme that persistently drives MAPK/ERK signaling independently of upstream RAS input. The kinase retains the typical regulatory and catalytic architecture of RAF proteins, including an N‑terminal RAS-binding and regulatory region and a C‑terminal kinase domain; the V600E change mimics phosphorylation within the activation segment, stabilizing the active conformation, increasing kinase activity by several hundred‑fold compared with wild-type BRAF, and favoring monomeric signaling that potently phosphorylates MEK1/2. Sustained MEK–ERK activation by BRAF V600E alters transcription of genes that control proliferation, differentiation and apoptosis and promotes melanocyte survival and cell-cycle progression through upregulation of cyclin D1, downregulation of p27^Kip1 and modulation of pro- and anti-apoptotic factors, while also impacting senescence programs and cooperating with additional lesions to overcome oncogene-induced growth arrest during melanomagenesis. BRAFV600E reshapes chromatin and gene expression patterns, with widespread changes in genes linked to cell adhesion, invasion, immune evasion and cytokine signaling, indicating that the mutation drives proliferation and also remodels the tumor microenvironment and metastatic behavior through MAPK-dependent transcriptional programs. In melanoma, activating BRAF mutations occur in about half of cases, and more than 90% of these involve V600E, defining a major molecular subset with characteristic clinicopathological features and strong dependence on MAPK signaling. BRAF V600E is also present in a distinct subgroup of colorectal cancers arising through the serrated pathway, where it appears early in tumorigenesis and associates with high-level microsatellite instability, CpG island methylator phenotype and right-sided colon location, and these tumors often exhibit poor prognosis and reduced benefit from standard anti-EGFR therapies due to MAPK pathway dominance. BRAF V600E in colorectal cancer intersects with WNT signaling, augmenting WNT activity in the absence of common APC mutations and relying on RNF43 and other alterations to sustain β‑catenin–driven transcription, highlighting cross-talk between MAPK and WNT cascades in this aggressive subtype. The central role of BRAF V600E in MAPK pathway activation has made it a key therapeutic target, and selective inhibitors such as vemurafenib and dabrafenib, often combined with MEK inhibitors, produce significant tumor regressions in BRAF V600-mutant melanoma and metastatic colorectal cancer, while acquired resistance frequently involves reactivation of MAPK signaling via secondary RAS/RAF alterations or activation of parallel pathways such as PI3K.

Background

BRAF V600E is an oncogenic variant of the RAF family serine/threonine kinase BRAF in which a valine-to-glutamate substitution at codon 600 within the activation segment of the kinase domain produces a constitutively active enzyme that persistently drives MAPK/ERK signaling independently of upstream RAS input. The kinase retains the typical regulatory and catalytic architecture of RAF proteins, including an N‑terminal RAS-binding and regulatory region and a C‑terminal kinase domain; the V600E change mimics phosphorylation within the activation segment, stabilizing the active conformation, increasing kinase activity by several hundred‑fold compared with wild-type BRAF, and favoring monomeric signaling that potently phosphorylates MEK1/2. Sustained MEK–ERK activation by BRAF V600E alters transcription of genes that control proliferation, differentiation and apoptosis and promotes melanocyte survival and cell-cycle progression through upregulation of cyclin D1, downregulation of p27^Kip1 and modulation of pro- and anti-apoptotic factors, while also impacting senescence programs and cooperating with additional lesions to overcome oncogene-induced growth arrest during melanomagenesis. BRAFV600E reshapes chromatin and gene expression patterns, with widespread changes in genes linked to cell adhesion, invasion, immune evasion and cytokine signaling, indicating that the mutation drives proliferation and also remodels the tumor microenvironment and metastatic behavior through MAPK-dependent transcriptional programs. In melanoma, activating BRAF mutations occur in about half of cases, and more than 90% of these involve V600E, defining a major molecular subset with characteristic clinicopathological features and strong dependence on MAPK signaling. BRAF V600E is also present in a distinct subgroup of colorectal cancers arising through the serrated pathway, where it appears early in tumorigenesis and associates with high-level microsatellite instability, CpG island methylator phenotype and right-sided colon location, and these tumors often exhibit poor prognosis and reduced benefit from standard anti-EGFR therapies due to MAPK pathway dominance. BRAF V600E in colorectal cancer intersects with WNT signaling, augmenting WNT activity in the absence of common APC mutations and relying on RNF43 and other alterations to sustain β‑catenin–driven transcription, highlighting cross-talk between MAPK and WNT cascades in this aggressive subtype. The central role of BRAF V600E in MAPK pathway activation has made it a key therapeutic target, and selective inhibitors such as vemurafenib and dabrafenib, often combined with MEK inhibitors, produce significant tumor regressions in BRAF V600-mutant melanoma and metastatic colorectal cancer, while acquired resistance frequently involves reactivation of MAPK signaling via secondary RAS/RAF alterations or activation of parallel pathways such as PI3K.

References
https://pubmed.ncbi.nlm.nih.gov/22554099/ https://pubmed.ncbi.nlm.nih.gov/30598662/

Protein Size (kDa)	PVDF Transfer Membrane Pore Size (μm)
< 10	0.22
10-20	0.22
20-30	0.45
30-45	0.45
45-70	0.45
80-100	0.45
100-140	0.45
> 140	0.45

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Protein Size (kDa)	Separating Gel Percentage
4-40	20%
12-45	15%
10-70	12.5%
15-100	10%
25-100	8%
60-210	5%