Overview

  • Product name
    Anti-BRAF (mutated V600 E) antibody [VE1]
    See all BRAF primary antibodies
  • Description
    Mouse monoclonal [VE1] to BRAF (mutated V600 E)
  • Host species
    Mouse
  • Specificity
    The VE1 monoclonal is a sensitive and specific antibody that detects mutated, constitutively active BRAF protein where glutamic acid is present at codon 600 instead of valine (V600E) (PubMed IDs: 21638088, 23657789).
  • Tested applications
    Suitable for: IHC-Pmore details
  • Species reactivity
    Reacts with: Human
    Predicted to work with: Mouse, Chicken
  • Immunogen

    Synthetic peptide within Human BRAF aa 550-650 (mutated V600 E). The exact sequence is proprietary.
    Database link: P15056

  • Positive control
    • IHC-P: Human colon carcinoma tissue with B-RAF V600E mutation.
  • General notes

    The V600E activating mutation in BRAF is found in several cancer types including: ~65% of pleomorphic xanthoastrocytomas, ~50% of melanomas, ~35% of papillary thyroid carcinomas and ~5% of microsatellite-stable colon cancers (PubMed IDs: 21274720, 24508103, 18682506, 16024606). 

    The majority (>90%) of BRAF mutant cancers harbor the V600E mutation. The mutation leads to activation of the MAPK signaling pathway that increases cell invasion and reduces apoptosis. It also leads to reduced expression of melanocyte differentiation antigens and subsequent immune evasion (PubMed IDs: 21638088, 20551059). 

    The VE1 monoclonal antibody is manufactured by Abcam. If you require a particular buffer formulation or a particular conjugate for your experiments, please contact orders@abcam.com or you can find further information here.

Properties

Applications

Our Abpromise guarantee covers the use of ab228461 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
IHC-P 1/100.

Target

  • Function
    Involved in the transduction of mitogenic signals from the cell membrane to the nucleus. May play a role in the postsynaptic responses of hippocampal neuron.
  • Tissue specificity
    Brain and testis.
  • Involvement in disease
    Note=Defects in BRAF are found in a wide range of cancers.
    Defects in BRAF may be a cause of colorectal cancer (CRC) [MIM:114500].
    Defects in BRAF are involved in lung cancer (LNCR) [MIM:211980].
    Defects in BRAF are involved in non-Hodgkin lymphoma (NHL) [MIM:605027]. NHL is a cancer that starts in cells of the lymph system, which is part of the body's immune system. NHLs can occur at any age and are often marked by enlarged lymph nodes, fever and weight loss.
    Defects in BRAF are a cause of cardiofaciocutaneous syndrome (CFC syndrome) [MIM:115150]; also known as cardio-facio-cutaneous syndrome. CFC syndrome is characterized by a distinctive facial appearance, heart defects and mental retardation. Heart defects include pulmonic stenosis, atrial septal defects and hypertrophic cardiomyopathy. Some affected individuals present with ectodermal abnormalities such as sparse, friable hair, hyperkeratotic skin lesions and a generalized ichthyosis-like condition. Typical facial features are similar to Noonan syndrome. They include high forehead with bitemporal constriction, hypoplastic supraorbital ridges, downslanting palpebral fissures, a depressed nasal bridge, and posteriorly angulated ears with prominent helices. The inheritance of CFC syndrome is autosomal dominant.
    Defects in BRAF are the cause of Noonan syndrome type 7 (NS7) [MIM:613706]. Noonan syndrome is a disorder characterized by facial dysmorphic features such as hypertelorism, a downward eyeslant and low-set posteriorly rotated ears. Other features can include short stature, a short neck with webbing or redundancy of skin, cardiac anomalies, deafness, motor delay and variable intellectual deficits.
    Defects in BRAF are the cause of LEOPARD syndrome type 3 (LEOPARD3) [MIM:613707]. LEOPARD3 is a disorder characterized by lentigines, electrocardiographic conduction abnormalities, ocular hypertelorism, pulmonic stenosis, abnormalities of genitalia, retardation of growth, and sensorineural deafness.
    Note=A chromosomal aberration involving BRAF is found in pilocytic astrocytomas. A tandem duplication of 2 Mb at 7q34 leads to the expression of a KIAA1549-BRAF fusion protein with a constitutive kinase activity and inducing cell transformation.
  • Sequence similarities
    Belongs to the protein kinase superfamily. TKL Ser/Thr protein kinase family. RAF subfamily.
    Contains 1 phorbol-ester/DAG-type zinc finger.
    Contains 1 protein kinase domain.
    Contains 1 RBD (Ras-binding) domain.
  • Cellular localization
    Nucleus. Cytoplasm. Cell membrane. Colocalizes with RGS14 and RAF1 in both the cytoplasm and membranes.
  • Information by UniProt
  • Database links
  • Alternative names
    • FLJ95109 antibody
    • 94 kDa B raf protein antibody
    • B raf 1 antibody
    • B raf antibody
    • B Raf proto oncogene serine threonine protein kinase antibody
    • B Raf proto oncogene, serine/threonine kinase antibody
    • B RAF1 antibody
    • B-Raf proto-oncogene serine/threonine-protein kinase (p94) antibody
    • BRAF 1 antibody
    • BRAF antibody
    • BRAF_HUMAN antibody
    • BRAF1 antibody
    • cRmil antibody
    • MGC126806 antibody
    • MGC138284 antibody
    • Murine sarcoma viral (v-raf) oncogene homolog B1 antibody
    • Murine sarcoma viral v raf oncogene homolog B1 antibody
    • NS7 antibody
    • Oncogen BRAF antibody
    • oncogene BRAF1 antibody
    • p94 antibody
    • Proto-oncogene B-Raf antibody
    • Proto-oncogene c-Rmil antibody
    • RAFB 1 antibody
    • RAFB1 antibody
    • RMIL antibody
    • Serine/threonine-protein kinase B-raf antibody
    • v raf murine sarcoma viral oncogene homolog B antibody
    • v raf murine sarcoma viral oncogene homolog B1 antibody
    • v-Raf murine sarcoma viral oncogene homolog B1 antibody
    see all

Images

  • Samples from human tumor tissues that were Positive (A, E, I) and Negative (B, F, J) for BRAF expression. Boxes in A, E, I show the negative controls from their corresponding non-tumor tissues. 

    IHC for BRAF protein expression was performed on 4 μm-thick sections of formalin-fixed, paraffin-embedded tissues, using ab228461. The specimens were fixed in 10% neutral buffered formalin for 24–48 hours. after deparaffinization, the slides were pretreated with cell conditioning 1 for 64 minutes for antigen unmasking and followed by pre-primary antibody peroxidase inhibition. The slides were then incubated with the VE1 antibody at 37°C for 16 minutes, and counterstained with hematoxylin II for 4 minutes and bluing reagent for 4 minutes.

  • BRAF (mutated V600E) VE1 immunoreactivity in melanoma sections: (A) strongly positive homogeneous staining, (B) moderately positive homogeneous staining, (C) weakly positive homogeneous staining and (D) negative staining of a patients biopsy that was identified as BRAF (mutated V600D).

     

    Sections were stained with undiluted ab228461. pre-treated with cell conditioner 1 (pH 8) for 60 min, followed by incubation with ab228461 at 37 °C for 32 min. Counterstained with one drop of haematoxylin for 4 min and one drop of bluing reagent for 4 min.

  • (A) Positive staining in a V600E2-mutated melanoma sample of patient 8 (homogenous brown colour indicating mutated cells) with ab228461 and (B) negative control.

  • BRAF V600E IHC positive PXA cases molecularly confirmed as BRAF V600E mutant tumors: strong granular cytoplasmic immunostaining of a characteristic pleomorphic multinucleated giant tumor cell (A), weak granular cytoplasmic immunostaining of pleomorphic and spindle tumor cells (B), strong granular cytoplasmic immunostaining of a cluster of isolated tumor cells (C).

     

    Four-micron freshly cut sections (<2 weeks) of formalin-fixed, paraffin-embedded (FFPE) tissue from human were dried and melted at 62°C oven for 20 minutes. Subsequently, they were stained with ab228461. Staining was performed on the Ventana BenchMark XT (Ventana Medical Systems Inc.). The staining protocol included online deparaffinization, HIER (Heat Induced Epitope Retrieval) with Ventana Cell Conditioning 1 for 32 minutes and primary antibody incubation for 32 minutes at 37°C. 

  • BRAF V600E mutations in adamantinomatous craniopharyngioma. Two cases are shown (a–e and f–j). a, f Classical features of aCP (wet keratin, stellate reticulum, palisaded epithelium). b, g Translocation of β-catenin is shown in both cases (brown reaction product). c, h An antibody to BRAF V600E (VE1) shows staining of the tumour tissue (brown reaction product).

    Immunohistochemical analysis was performed on 4µm sections from FFPE tumour specimens. Following dewaxing through graded alcohols, endogenous peroxidase activity was blocked (3% (v/v) H2O2 in PBS, pH 7.3, 20 minutes with orbital shaking). Epitope retrieval was achieved by autoclaving in sodium citrate (10mM, pH 6.0, 10 minutes) (β-catenin) or Tris-EDTA (10mM Tris base, 1mM EDTA, pH 9.0, 10 minutes) (BRAF). Sections were blocked with serum block for 15 minutes, then incubated overnight at 4°C with primary antibody diluted in PBS or TBS-T (1:1000 in PBS (β-catenin); 1:50 in TBS-T (BRAF)).

  • Representative photomicrographs serially stained for hematoxylin and eosin, PMS2 and BRAFV600E of (a) DNA mismatch repair-proficient/BRAF wild-type colorectal carcinoma and (b) DNA mismatch repair-proficient/BRAFV600E mutant tumor.

    The slides were dewaxed in Bond Dewax solution and hydrated in Bond Wash solution. Heat-induced epitope retrieval was performed for 60 min in the manufacturer’s alkaline retrieval solution ER2. Slides were then incubated with the primary antibody at a dilution of 1 in 80 for 30 min at room temperature. Antibody detection was performed using the biotin-free Bond Polymer Defined Detection System. Slides were then counterstained with hematoxylin.

     

  • Formalin-fixed, paraffin-embedded human colon carcinoma tissue stained for B Raf (mutated V600E) using ab228461 at a 1/100 dilution in immunohistochemical analysis.

References

This product has been referenced in:
  • Pagès M  et al. Co-occurrence of histone H3 K27M and BRAF V600E mutations in paediatric midline grade I ganglioglioma. Brain Pathol 28:103-111 (2018). Read more (PubMed: 27984673) »
  • Yoshimoto K  et al. High-resolution melting and immunohistochemical analysis efficiently detects mutually exclusive genetic alterations of adamantinomatous and papillary craniopharyngiomas. Neuropathology 38:3-10 (2018). Read more (PubMed: 28840946) »
See all 88 Publications for this product

Customer reviews and Q&As

Abcam has not validated the combination of species/application used in this Abreview.
Application
Western blot
Sample
Human Cell lysate - whole cell (CHL-1 and MEL-624)
Gel Running Conditions
Reduced Denaturing (4-12% Bis-Tris)
Loading amount
10 µg
Treatment
with varying (0, 0.1, 0.3, 1, 3, 10 µM) B-RAF and MEK inhibitors for 48 hours
Specification
CHL-1 and MEL-624
Blocking step
Milk as blocking agent for 30 minute(s) · Concentration: 5% · Temperature: 25°C

Allen Zhu

Verified customer

Submitted Jul 31 2018

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"

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