Overview

  • Product name
    Recombinant human B Raf + Cdc37 protein
  • Protein length
    Protein fragment

Description

  • Nature
    Recombinant
  • Source
    Baculovirus infected Sf9 cells
  • Amino Acid Sequence
    • Species
      Human
    • Sequence
      SEDRNRMKT LGRRDSSDDW EIPDGQITVG QRIGSGSFGT VYKGKWHGDV AVKMLNVTAP TPQQLQAFKN EVGVLRKTRH VNILLFMGYS TKPQLAIVTQ WCEGSSLYHH LHIIETKFEM IKLIDIARQT AQGMDYLHAK SIIHRDLKSN NIFLHEDLTV KIGDFGLATV KSRWSGSHQF EQLSGSILWM APEVIRMQDK NPYSFQSDVY AFGIVLYELM TGQLPYSNIN NRDQIIFMVG RGYLSPDLSK VRSNCPKAMK RLMAECLKKK RDERPLFPQI LASIELLARS LPKIH
    • Tags
      His tag N-Terminus , DDDDK tag N-Terminus
    • Additional sequence information
      Human BRAF (NM_004333, a.a.432-725, N-term His-tag, 36 kDa), and p50 (NM_007065, a.a.2-378 (end), N-term FLAG-tag, 45 kDa). Protein exists in a complex with Cdc37 and p50.

Associated products

Specifications

Our Abpromise guarantee covers the use of ab42596 in the following tested applications.

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

  • Biological activity

    Specific Activity: 12 pmol/min/μg.

    Activity was tested using 1 μM MEK1 (K97R) as substrate. Assay was performed in a 50 μl reation in 10 mM Tris, pH 7.4, 5 mM

    MgCl2 , 0.5 mM EDTA, 0.5 mM DTT, 5 μM ATP, 5 μg MEK1 (K97R) as substrate, and B Raf. 62 ng of B Raf enzyme phosphorylated over 18% of 1 μM MEK1 at room temperature in 1 hr.

  • Applications

    Inhibition Assay

  • Form
    Liquid
  • Additional notes

    Coexpressed in Baculovirus infected Sf9 cell expression system.

    Used for the study of enzyme kinetics, screening inhibitors, and selectivity profiling.

     

  • Concentration information loading...

Preparation and Storage

  • Stability and Storage

    Shipped on Dry Ice. Upon delivery aliquot. Store at -80°C. Avoid freeze / thaw cycle.

    pH: 8.00
    Preservative: 1.53% Imidazole
    Constituents: 0.018% Potassium chloride, 0.0462% DTT, 0.7% Tris HCl, 10% Glycerol, 0.72% Sodium chloride

    This product is an active protein and may elicit a biological response in vivo, handle with caution.

General Info

  • Cellular localization
    B Raf: Nucleus. Cytoplasm. Cell membrane. Colocalizes with RGS14 and RAF1 in both the cytoplasm and membranes. Cdc37: Cytoplasm.

Images

  • 10% SDS-PAGE with coomassie staining.  

    Left lane: 4 µg B Raf

    Right lane: Protein marker

References

ab42596 has not yet been referenced specifically in any publications.

Customer reviews and Q&As

1-4 of 4 Abreviews or Q&A

Answer

Thanks for your reply, and I'm happy to hear that ab55690 will work as a replacement.

I'm sending a free of charge vial on the order ***, which is on back order but is expected to arrive on Wednesday, March 7.

Please keep me updated about the results using this protein, and let me know if you have any questions or need anything else from me.

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Answer

Thank you for your reply and for your patience. I forwarded the assay details to the lab to see if there was any explanation for these results.

I have been informedthat there is 250 mM imidazole in the His-BRaf formulation. Imidazoleinterferes with anti-His antibody binding to the tag, which explains the resultsof the inhibition assay.





My contact at the lab recommendedtrying anADP glo kinase assay which uses MEK1 as a substrate, or you could remove the imidazole by dialysisor some other meansfor the TR-FRET assay.

As the ab42596 datasheet did not state that the protein contained imidazole, I would also be happy to send a replacement product or issue a credit or refund for your purchase of this protein. We have another B Raf protein, ab55690, except it is not His-tagged.

I look forward to hearing from you. I apologize for this inconvenience but I'm hopeful that we can resolve this issue promptly. Please let me know if you have any questions or if there is anything else that we can do for you.

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Answer

Thank you for contactin us and letting us know about this issue with ab42596.

I am sorry to hear about the trouble with this protein, andour files indicatethat you have previously contacted us about a delay in shipping this order to you.

In order to better understand this situation, I do have a couple additional questions:

1) Have you confirmed that the anti-His antibody is recognizing the His-tagged B Raf through any other assay?
2) Could you send a few details about the protocol (i.e. concentrations of the reagents in each step) for our records?
3) Do you have any data that you would be able to send?

We do guarantee all of our products to work as stated on the datasheet for up to 6 months after purchase, and this protein is guaranteed to work in inhibitiion assays, so I would be happy to send a replacement product or issue a credit or refund if you prefer.

I look forward to hearing from you. Please let me know if you have any additional questions or if there is anything else that we can do for you.

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Answer

Thank you for contacting Abcam regarding ab42595.

I am sorry that you have been experiencing difficulties with this protein. I have checked into the shipping error you referred to, and it seems that there was a delay in getting this shipment from our UK office, however conditions were such that the product quality was not compromised.

I would like to get some additional information regarding how you were using the product to see if there are any suggestions I could make to help improve your results. Can you please provide a description of your protocol and the results you obtained?

I look forward to your reply so that I may assist you further. Please do not hesitate to contact me if you have any additional questions.

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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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