Human BTK Antibody (566928)

R&D Systems | Catalog # MAB5807

R&D Systems

Key Product Details

Species Reactivity

Human

Applications

Western Blot, Immunocytochemistry, Simple Western

Label

Unconjugated

Antibody Source

Monoclonal Mouse IgG1 Clone # 566928
View all BTK Primary Antibodies »

Product Specifications

Immunogen

E. coli-derived recombinant human BTK
Glu280-Val377
Accession # Q06187

Specificity

Detects endogenous human BTK in Western blots.

Clonality

Monoclonal

Host

Mouse

Isotype

IgG1

Scientific Data Images for Human BTK Antibody (566928)

Detection of Human BTK by Western Blot.

Western Blot shows lysates of K562 human chronic myelogenous leukemia cell line. PVDF membrane was probed with 0.05 µg/ml of Mouse Anti-Human BTK Monoclonal Antibody (Catalog # MAB5807) followed by HRP-conjugated Anti-Mouse IgG Secondary Antibody (Catalog # HAF018). A specific band was detected for BTK at approximately 75 kDa (as indicated). This experiment was conducted under reducing conditions and using Western Blot Buffer Group 1.
BTK antibody in Raji Human Cell Line by Immunocytochemistry (ICC).

BTK in Raji Human Cell Line.

BTK was detected in immersion fixed Raji human Burkitt's lymphoma cell line using Mouse Anti-Human BTK Monoclonal Antibody (Catalog # MAB5807) at 10 µg/mL for 3 hours at room temperature. Cells were stained using the NorthernLights™ 557-conjugated Anti-Mouse IgG Secondary Antibody (red, upper panel; NL007) and counterstained with DAPI (blue, lower panel). Specific staining was localized to cytoplasm. View our protocol for Fluorescent ICC Staining of Non-adherent Cells.

Detection of Human BTK by Simple WesternTM.

Simple Western shows lysates of Daudi human Burkitt's lymphoma cell line and K562 human chronic myelogenous leukemia cell line, loaded at 0.2 mg/ml. A specific band was detected for BTK at approximately 75 kDa (as indicated) using 10 µg/mL of Mouse Anti-Human BTK Monoclonal Antibody (Catalog # MAB5807). This experiment was conducted under reducing conditions and using the 12‑230 kDa separation system.
Detection of BTK by Flow Cytometry

Detection of BTK by Flow Cytometry

Optimization of the AAV6–co-BTK donor cassette in DG75 BTK KO cells. A, A representation of the BTK locus is shown, including the sequence of the therapeutic gRNA targeting exon 2 and the gRNA used to knockout BTK by mutating exon 3. Arrows show the approximate cut site of both gRNAs (left). DG75 cells were edited with a Cas9/gRNA RNP targeting exon 3 of the BTK locus before deriving a clonal KO population (DG75 KO). Flow cytometry analysis of BTK expression in WT and KO DG75 cells (right). B, DG75 BTK KO cells were edited with a Cas9/gRNA RNP complex targeting exon 2 and transduced with an AAV6 HDR donor template containing a codon optimized BTK coding sequence, before deriving clonal populations. Western blot analysis of BTK levels in WT, KO, and a BTK “knocked in” clone. C, Real-time quantitative PCR evaluation of BTK mRNA expression in clones knocked in with a BTK codon-optimized coding sequence (CO; black dots) and a BTK codon-optimized coding sequence containing a canonical Kozak sequence alone (K; each clone shown with different color) or with a 300-bp region from I18 (I18; each clone with different color). The 2− delta delta CT values [Delta Ct single clones (Ct coBTK-Ct GAPDH)-deltaCt WT (Ct wtBTK-Ct GAPDH)] are shown. Of note, primers to amplify the WT and codon optimized BTK differ, so conclusions on the relative amount of BTK versus WT cannot be drawn. (Shown is the mean ± SD; ∗P <.005 1-way ANOVA with Tukey’s multiple comparison.) NS, Not significant; SSC, side scatter. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/38479630), licensed under a CC-BY license. Not internally tested by R&D Systems.
Detection of BTK by Western Blot

Detection of BTK by Western Blot

Optimization of a gene editing platform for XLA. A, Schematic of the gene editing strategy to knock in a BTK coding sequence into exon 2 of the native locus using the CRISPR/Cas9 system and an AAV6 to deliver the HDR donor template. The HDR donor template consists of a codon optimized version of the BTK coding sequence (co-BTK) with a canonical GCCACCATGG Kozak sequence (K), a 300 bp region of intron 18 (I18) and a beta growth hormone polyadenylation signal (poly A), flanked by the homology arms left (HAL) and right (HAR). B, Western blot analysis of BTK expression in DG75 and Jurkat T-cell clones. DG75 and Jurkat T cells were electroporated with the Cas9/gRNA RNP and transduced with the AAV6 vector containing the co-BTK cassette before clones were derived. The GAPDH housekeeping protein is used as a loading control. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/38479630), licensed under a CC-BY license. Not internally tested by R&D Systems.

Applications for Human BTK Antibody (566928)

Application
Recommended Usage

Immunocytochemistry

8-25 µg/mL
Sample: Immersion fixed Raji human Burkitt's lymphoma cell line

Simple Western

10 µg/mL
Sample: Daudi human Burkitt's lymphoma cell line and K562 human chronic myelogenous leukemia cell line

Western Blot

0.05 µg/mL
Sample: K562 human chronic myelogenous leukemia cell line

Reviewed Applications

Read 1 review rated 5 using MAB5807 in the following applications:

Formulation, Preparation, and Storage

Purification

Protein A or G purified from hybridoma culture supernatant

Reconstitution

Reconstitute at 0.5 mg/mL in sterile PBS. For liquid material, refer to CoA for concentration.

Formulation

Lyophilized from a 0.2 μm filtered solution in PBS with Trehalose. See Certificate of Analysis for details.<br/>*Small pack size (-SP) is supplied either lyophilized or as a 0.2 µm filtered solution in PBS.

Shipping

Lyophilized product is shipped at ambient temperature. Liquid small pack size (-SP) is shipped with polar packs. Upon receipt, store immediately at the temperature recommended below.

Stability & Storage

Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
  • 12 months from date of receipt, -20 to -70 °C as supplied.
  • 1 month, 2 to 8 °C under sterile conditions after reconstitution.
  • 6 months, -20 to -70 °C under sterile conditions after reconstitution.

Calculators

The reconstitution calculator allows you to quickly calculate the volume of a reagent to reconstitute your vial. Simply enter the mass of reagent and the target concentration and the calculator will determine the rest.

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Background: BTK

Bruton tyrosine kinase (BTK) is a 75 kDa cytoplasmic protein tyrosine kinase that is widely expressed in hematopoietic cells. BTK is required for B cell receptor signaling and B cell development. Defects in BTK result in X-linked agammaglobulinemia which is characterized by a severely decreased level of circulating antibodies. Like other Tec family kinases, BTK contains a Pleckstrin homology domain, a Tec homology domain, an SH3 domain, an SH2 domain, and a protein kinase domain. Association of the BTK SH2 domain with the B cell linker protein (BLNK) is required for the activation of PLC gamma by BTK. Within the SH2 domain (aa 280‑377), human BTK shares 99% aa sequence identity with mouse and rat BTK.

Long Name

Bruton Agammaglobulinemia Tyrosine Kinase

Alternate Names

AGMX1, IMD1, PSCTK1, XLA

Entrez Gene IDs

695 (Human)

Gene Symbol

BTK

UniProt

Q06187

Additional BTK Products

Product Documents for Human BTK Antibody (566928)

Certificate of Analysis

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Note: Certificate of Analysis not available for kit components.

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Product Specific Notices for Human BTK Antibody (566928)

For research use only

Citations for Human BTK Antibody (566928)

Customer Reviews for Human BTK Antibody (566928) (1)

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  • Human BTK Antibody
    Name: Anonymous
    Application: Immunocytochemistry/Immunofluorescence
    Sample Tested: Jurkat human acute T cell leukemia cell line
    Species: Human
    Verified Customer | Posted 07/07/2022
    Human BTK Antibody (566928) MAB5807

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Protocols

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