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RecombinantRabMAb

Recombinant Anti-BTK antibody [EPR20445] - BSA and Azide free (ab227812)

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-BTK antibody [EPR20445] - BSA and Azide free (ab227812)
  • Western blot - Anti-BTK antibody [EPR20445] - BSA and Azide free (ab227812)
  • Immunocytochemistry/ Immunofluorescence - Anti-BTK antibody [EPR20445] - BSA and Azide free (ab227812)
  • Flow Cytometry (Intracellular) - Anti-BTK antibody [EPR20445] - BSA and Azide free (ab227812)
  • Western blot - Anti-BTK antibody [EPR20445] - BSA and Azide free (ab227812)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-BTK antibody [EPR20445] - BSA and Azide free (ab227812)
  • Immunoprecipitation - Anti-BTK antibody [EPR20445] - BSA and Azide free (ab227812)
  • Anti-BTK antibody [EPR20445] - BSA and Azide free (ab227812)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EPR20445] to BTK - BSA and Azide free
  • Suitable for: Flow Cyt (Intra), WB, IHC-P, ICC/IF, IP
  • Reacts with: Mouse, Rat, Human

Conjugates logo Related conjugates and formulations

Unconjugated

Overview

  • Product name

    Anti-BTK antibody [EPR20445] - BSA and Azide free
    See all BTK primary antibodies

  • Description

    Rabbit monoclonal [EPR20445] to BTK - BSA and Azide free

  • Host species

    Rabbit

  • Specificity

    This antibody does not react with mouse and rat species in Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) application.

  • Tested applications

    Suitable for: Flow Cyt (Intra), WB, IHC-P, ICC/IF, IPmore details

  • Species reactivity

    Reacts with: Mouse, Rat, Human

  • Immunogen

    Recombinant fragment. This information is proprietary to Abcam and/or its suppliers.

  • Positive control

    • IHC-P: Human tonsil and colon tissues. ICC/IF: Ramos cells. Flow Cyt (intra): Ramos cells. IP: Ramos whole cell lysate. WB: THP-1, RAW 264.7 and NR8383 whole cell lysates.

  • General notes

    ab227812 is the carrier-free version of ab208937.

    Our carrier-free antibodies are typically supplied in a PBS-only formulation, purified and free of BSA, sodium azide and glycerol. The carrier-free buffer and high concentration allow for increased conjugation efficiency.

    This conjugation-ready format is designed for use with fluorochromes, metal isotopes, oligonucleotides, and enzymes, which makes them ideal for antibody labelling, functional and cell-based assays, flow-based assays (e.g. mass cytometry) and Multiplex Imaging applications.

    Use our conjugation kits for antibody conjugates that are ready-to-use in as little as 20 minutes with <1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.

    This product is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm, without the need for antibody preparation. Maxpar® is a trademark of Fluidigm Canada Inc.

    This product is a recombinant monoclonal antibody, which offers several advantages including:

    • - High batch-to-batch consistency and reproducibility
    • - Improved sensitivity and specificity
    • - Long-term security of supply
    • - Animal-free production
    For more information see here.

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

Properties

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab227812 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
Flow Cyt (Intra)
Use at an assay dependent concentration.
WB
Use at an assay dependent concentration. Detects a band of approximately 76 kDa (predicted molecular weight: 76 kDa).
IHC-P
Use at an assay dependent concentration. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
ICC/IF
Use at an assay dependent concentration.
IP
Use at an assay dependent concentration.
Notes
Flow Cyt (Intra)
Use at an assay dependent concentration.
WB
Use at an assay dependent concentration. Detects a band of approximately 76 kDa (predicted molecular weight: 76 kDa).
IHC-P
Use at an assay dependent concentration. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
ICC/IF
Use at an assay dependent concentration.
IP
Use at an assay dependent concentration.

Target

  • Function

    Plays a crucial role in B-cell ontogeny. Transiently phosphorylates GTF2I on tyrosine residues in response to B-cell receptor cross-linking. Required for the formation of functional ARID3A DNA-binding complexes.

  • Involvement in disease

    Defects in BTK are the cause of X-linked agammaglobulinemia (XLA) [MIM:300755]; also known as X-linked agammaglobulinemia type 1 (AGMX1) or immunodeficiency type 1 (IMD1). XLA is a humoral immunodeficiency disease which results in developmental defects in the maturation pathway of B-cells. Affected boys have normal levels of pre-B-cells in their bone marrow but virtually no circulating mature B-lymphocytes. This results in a lack of immunoglobulins of all classes and leads to recurrent bacterial infections like otitis, conjunctivitis, dermatitis, sinusitis in the first few years of life, or even some patients present overwhelming sepsis or meningitis, resulting in death in a few hours. Treatment in most cases is by infusion of intravenous immunoglobulin.
    Defects in BTK may be the cause of X-linked hypogammaglobulinemia and isolated growth hormone deficiency (XLA-IGHD) [MIM:307200]; also known as agammaglobulinemia and isolated growth hormone deficiency or Fleisher syndrome or isolated growth hormone deficiency type 3 (IGHD3). In rare cases XLA is inherited together with isolated growth hormone deficiency (IGHD).

  • Sequence similarities

    Belongs to the protein kinase superfamily. Tyr protein kinase family. TEC subfamily.
    Contains 1 Btk-type zinc finger.
    Contains 1 PH domain.
    Contains 1 protein kinase domain.
    Contains 1 SH2 domain.
    Contains 1 SH3 domain.

  • Post-translational
    modifications

    Autophosphorylated on Tyr-223 and Tyr-551. Phosphorylation of Tyr-223 may create a docking site for a SH2 containing protein.

  • Cellular localization

    Cytoplasm. Membrane. Nucleus.
  • Information by UniProt

  • Database links

    • Alternative names

      • Agammaglobulinaemia tyrosine kinase antibody
      • AGMX 1 antibody
      • AGMX1 antibody
      • AT antibody
      • ATK antibody
      • B cell progenitor kinase antibody
      • B-cell progenitor kinase antibody
      • BPK antibody
      • Bruton agammaglobulinemia tyrosine kinase antibody
      • Bruton tyrosine kinase antibody
      • Bruton’s Tyrosine Kinase antibody
      • Btk antibody
      • BTK_HUMAN antibody
      • dominant-negative kinase-deficient Brutons tyrosine kinase antibody
      • IMD 1 antibody
      • IMD1 antibody
      • MGC126261 antibody
      • MGC126262 antibody
      • OTTHUMP00000063593 antibody
      • PSCTK 1 antibody
      • PSCTK1 antibody
      • truncated Bruton agammaglobulinemia tyrosine kinase antibody
      • Tyrosine protein kinase BTK antibody
      • Tyrosine-protein kinase BTK antibody
      • tyrosine-protein kinase BTK isoform (lacking exon 14 antibody
      • XLA antibody
      see all

    Images

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-BTK antibody [EPR20445] - BSA and Azide free (ab227812)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-BTK antibody [EPR20445] - BSA and Azide free (ab227812)

      Immunohistochemical analysis of paraffin-embedded human colon tissue labeling BTK with ab208937 at 1/4000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Strong cytoplasmic staining on lymphoid nodule in human colon (PMID: 25433814). Counter stained with hematoxylin.

      Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab208937).

      Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

    • Western blot - Anti-BTK antibody [EPR20445] - BSA and Azide free (ab227812)
      Western blot - Anti-BTK antibody [EPR20445] - BSA and Azide free (ab227812)
      All lanes : Anti-BTK antibody [EPR20445] (ab208937) at 1/1000 dilution

      Lane 1 : THP-1 (human monocytic leukemia cell) whole cell lysate
      Lane 2 : RAW 264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysate
      Lane 3 : NR8383 (rat lung macrophage (alveolar)) whole cell lysate

      Lysates/proteins at 20 µg per lane.

      Secondary
      All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

      Predicted band size: 76 kDa
      Observed band size: 76 kDa


      Exposure time: 37 seconds


      Blocking buffer and concentration: 5% NFDM/TBST
      Diluting buffer and concentration: 5% NFDM/TBST

      Lysates were freshly made and used for Western blotting immediately to minimize protein degradation.

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab208937).

    • Immunocytochemistry/ Immunofluorescence - Anti-BTK antibody [EPR20445] - BSA and Azide free (ab227812)
      Immunocytochemistry/ Immunofluorescence - Anti-BTK antibody [EPR20445] - BSA and Azide free (ab227812)

      Immunofluorescent analysis of 100% methanol-fixed Ramos (human Burkitt's lymphoma cell line) and Jurkat (human T cell leukemia cell line from peripheral blood) cells labeling BTK with ab208937 at 1/100 dilution followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

      Confocal image showing cytoplasmic staining on Ramos cell line.

      Negative control: Jurkat cell line (PMID: 24759210).

      The nuclear counter stain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) (ab195889) (red) at 1/200 dilution.

      Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution.

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab208937).

    • Flow Cytometry (Intracellular) - Anti-BTK antibody [EPR20445] - BSA and Azide free (ab227812)
      Flow Cytometry (Intracellular) - Anti-BTK antibody [EPR20445] - BSA and Azide free (ab227812)

      Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol-permeabilized  Jurkat (human T cell leukemia cell line from peripheral blood) (left panel) and Ramos (human Burkitt's lymphoma cell line) (right panel) cell lines labeling  BTK with ab208937 at 1/600 dilution (red) compared with a Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730) (black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) at 1/2000 dilution was used as the secondary antibody. 

      Negative control: Jurkat cell line (PMID: 24759210). 

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab208937). 

       

       

    • Western blot - Anti-BTK antibody [EPR20445] - BSA and Azide free (ab227812)
      Western blot - Anti-BTK antibody [EPR20445] - BSA and Azide free (ab227812)
      All lanes : Anti-BTK antibody [EPR20445] (ab208937) at 1/1000 dilution

      Lane 1 : RAW 264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysate
      Lane 2 : NIH/3T3 (mouse embryonic fibroblast) whole cell lysate

      Lysates/proteins at 20 µg per lane.

      Secondary
      All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

      Predicted band size: 76 kDa
      Observed band size: 76 kDa


      Exposure time: 15 seconds


      Blocking buffer and concentration: 5% NFDM/TBST
      Diluting buffer and concentration: 5% NFDM/TBST

      Lysates were freshly made and used for Western blotting immediately to minimize protein degradation.

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab208937).

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-BTK antibody [EPR20445] - BSA and Azide free (ab227812)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-BTK antibody [EPR20445] - BSA and Azide free (ab227812)

      Immunohistochemical analysis of paraffin-embedded human tonsil tissue labeling BTK with ab208937 at 1/4000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Strong cytoplasmic staining in mantle zone and weaker cytoplasmic staining in the germinal center of human tonsil (PMID: 25433814). Counter stained with hematoxylin.

      Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab208937).

      Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

    • Immunoprecipitation - Anti-BTK antibody [EPR20445] - BSA and Azide free (ab227812)
      Immunoprecipitation - Anti-BTK antibody [EPR20445] - BSA and Azide free (ab227812)

      BTK was immunoprecipitated from 0.35 mg of Ramos (human Burkitt's lymphoma cell line) whole cell lysate with ab208937 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab208937 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.

      Lane 1: Ramos whole cell lysate 10 µg (Input). 

      Lane 2: ab208937 IP in Ramos whole cell lysate. 

      Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab208937 in Ramos whole cell lysate.

      Blocking and dilution buffer and concentration: 5% NFDM/TBST.

      Exposure time: 3 seconds.

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab208937).

    • Anti-BTK antibody [EPR20445] - BSA and Azide free (ab227812)
      Anti-BTK antibody [EPR20445] - BSA and Azide free (ab227812)

    Protocols

    To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.

    Click here to view the general protocols

    References (0)

    Publishing research using ab227812? Please let us know so that we can cite the reference in this datasheet.

    ab227812 has not yet been referenced specifically in any publications.

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