Overview

  • Product name
    Anti-BRAF antibody [EP152Y]
    See all BRAF primary antibodies
  • Description
    Rabbit monoclonal [EP152Y] to BRAF
  • Host species
    Rabbit
  • Specificity

    The mouse and rat recommendation is based on the WB results. We do not guarantee IHC-P for mouse and rat.

     

  • Tested applications
    Suitable for: WB, IHC-P, IPmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide within Human BRAF aa 50-150. The exact sequence is proprietary.
    Database link: P15056

  • Positive control
    • WB: HeLa whole cell lysate (ab150035), PC-12 whole cell lysate, mouse brain tissue lysate IHC-P: human prostate cancer tissue, Human breast cancer tissue. This antibody also reacts with rat brain tissue. IP: HeLa cells.
  • General notes

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMab® patents.

    We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

    This product is a recombinant rabbit monoclonal antibody.

Properties

Applications

Our Abpromise guarantee covers the use of ab33899 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
WB 1/1000 - 1/5000. Detects a band of approximately 87 kDa (predicted molecular weight: 85 kDa).
IHC-P 1/100.

See IHC antigen retrieval protocols.

The mouse and rat recommendation is based on the WB results. We do not guarantee IHC-P for mouse and rat.

IP 1/50.

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

  • Lane 1: Wild-type HAP1 whole cell lysate (40 µg)
    Lane 2: B Raf knockout HAP1 whole cell lysate (40 µg)
    Lane 3: HeLa whole cell lysate (40 µg)

    Lanes 1 - 3: Merged signal (red and green). Green - ab33899 (unpurified) observed at 90 kDa. Red - loading control, ab8245, observed at 37 kDa.

    ab33899 was shown to recognize B Raf in wild-type HAP1 cells as signal was lost at the expected MW in B Raf knockout cells. Additional cross-reactive bands were observed in the wild-type and knockout cells. Wild-type and B Raf knockout samples were subjected to SDS-PAGE. ab33899 and ab8245 (Mouse anti-GAPDH loading control) were incubated overnight at 4°C at 1/1000 dilution and 1/20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed ab216776 secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Human breast cancer tissue sections labeling BRAF with purified ab33899 at 1/100 dilution (4.77 µg/ml). Heat mediated antigen retrieval was performed using heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0). ImmunoHistoProbe one step HRP Polymer (ready to use) was used as the secondary antibody. Negative control: PBS instead of the primary antibody. Hematoxylin was used as a counterstain.

  • Anti-BRAF antibody [EP152Y] (ab33899) at 1/5000 dilution (Purified) + PC-12 (Rat adrenal gland pheochromocytoma) whole cell lysates at 15 µg

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

    Predicted band size: 85 kDa
    Observed band size: 87 kDa
    why is the actual band size different from the predicted?

  • All lanes : Anti-BRAF antibody [EP152Y] (ab33899) at 1/2000 dilution (Purified)

    Lane 1 : HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysates
    Lane 2 : Mouse brain lysates

    Lysates/proteins at 15 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

    Predicted band size: 85 kDa
    Observed band size: 87 kDa why is the actual band size different from the predicted?

  • ab33899 (Purified) at 1:500 dilution (0.954 µg/ml) immunoprecipitating BRAF in HeLa whole cell lysate.
    Lane 1 (input): HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate 10 µg
    Lane 2 (+): ab33899 & HeLa whole cell lysate
    Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab33899 in HeLa whole cell lysate
    For western blotting, VeriBlot for IP secondary antibody (HRP) (ab131366) was used as the secondary antibody at 1:1000 dilution.
    Blocking and diluting buffer: 5% NFDM /TBST .
  • This image shows paraffin embedded human prostate cancer tissue sample stained with ab33899 (unpurified) at 1/250 dilution.

  • ab33899 (unpurified) staining B Raf cells from human prostate tissue by immunohistochemistry (formalin/PFA-fixed paraffin-embedded sections). Cells were formaldehyde fixed and permeabilized in PBS-Tween 20 prior to blocking in 70% serum for 10 minutes at 25°C. The primary antibody was diluted 1/250 and incubated with the sample for 1 hour at 25°C. A biotin conjugated goat polyclonal to mouse Ig was used as the secondary.

    See Abreview

  • Anti-BRAF antibody [EP152Y] (ab33899) at 1/5000 dilution (unpurified) + HeLa cell lysate

    Predicted band size: 85 kDa
    Observed band size: 87 kDa why is the actual band size different from the predicted?

  • All lanes : Anti-BRAF antibody [EP152Y] (ab33899) at 1/1000 dilution (unpurified)

    Lane 1 : Lysate prepared from mouse brain
    Lane 2 : Lysate prepared from mouse heart
    Lane 3 : Lysate prepared from mouse kidney
    Lane 4 : Lysate prepared from mouse spleen
    Lane 5 : Lysate prepared from rat brain
    Lane 6 : Lysate prepared from rat heart
    Lane 7 : Lysate prepared from rat kidney
    Lane 8 : Lysate prepared from rat spleen

    Predicted band size: 85 kDa


    Exposure time: 3 minutes

References

This product has been referenced in:
  • Du Z  et al. Genome-wide transcriptional analysis of BRD4-regulated genes and pathways in human glioma U251 cells. Int J Oncol N/A:N/A (2018). Read more (PubMed: 29568956) »
  • Gu HJ & Zhou B Focal adhesion kinase promotes progression and predicts poor clinical outcomes in patients with osteosarcoma. Oncol Lett 15:6225-6232 (2018). Read more (PubMed: 29849782) »
See all 16 Publications for this product

Customer reviews and Q&As

1-5 of 5 Q&A

Question
Answer

Thank you for confirming these details and for your cooperation. The details provided enable us to closely monitor the quality of our products. I am sorry this product did not perform as stated on the datasheet and for the inconvenience this has caused. As requested, I have issued a free of charge replacement with the order number xxxxx. To check the status of the order please contact our Customer Service team and reference this number. Please note that this free of charge replacement vial is also covered by our Abpromise guarantee. Should you still be experiencing difficulties, or if you have any further questions, please do not hesitate to let us know. As we had spoken about, I have also created a discount code for use with . The code is: xxxx and is valid until: xxxx. I wish you the best of luck with your research.

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Question
Answer

Unfortunately shipment of this order is being held up by our accounts team.

Our accounting department can be contacted by email at mailto:us.credits@abcam.com or by telephone using the information at the Contact Us link in the top right corner of our website. Please refer to theorder reference numberin any correspondence with our accounting department.

I hope this is helpful. Please contact me again with any technical questions you may have.

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Answer

Thank you for your enquiry. I have been in touch with the lab and I can tell you that this antibody has NOT been tested for its capacity to differentiate between wild type and variant (V600E) BRAF. I could not find literature relating to the variant sequence of BRAF. However, I can tell you that the immunogen sequence used to raise this antibody was CGEHNPPSIYLEAYEEYT, corresponding to N terminal amino acids 70-86. I hope this information helps, please do not hesitate to contact us if you need any more advice or information.

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Answer

Thank you for your enquiry and patience. Ab33899 (B-raf antibody) was tested by Western Blotting on various mouse tissues (brain, heart, kidney, muscle) and was found to not cross-react with these tissues. This antibody should work fine in your system, human transgene in a mouse model.

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Answer

A synthetic peptide corresponding to residues near N-terminus of human B-Raf (aa-70-86) was used as an immunogen for anti-B-Raf Rabbit Monoclonal antibody ab33899. The exact peptide sequence is following: CGEHNPPSIYLEAYEEYT. I hope this information helps you,

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

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