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Human Bruton'S Tyrosine Kinase (Btk) ELISA Kit

Due to the possibility of mismatching between antigens from other origin and antibodies used in our kits (e.g., antibody targets conformational epitope rather than linear epitope), some native or recombinant proteins from other manufacturers may not be recognized by our products.

Principle of the Assay

The microtiter plate provided in this kit has been pre-coated with an antibody specific to Btk. Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated antibody preparation specific to Btk. Next, Avidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated. After the TMB substrate solution is added, only those wells that contain Btk, biotin-conjugated antibody, and enzyme-conjugated Avidin will exhibit a change in color. The enzyme-substrate reaction is terminated by the addition of sulphuric acid solution, and the color change is measured spectrophotometrically at a wavelength of 450nm ± 10nm. The concentration of Btk in the samples is then determined by comparing the O.D. of the samples to the standard curve.


For Use with serum, plasma, and cell culture supernatants. For Research Use Only. Not for use in diagnostic procedures.

Target Information

Non-receptor tyrosine kinase indispensable for B lymphocyte development, differentiation and signaling. Binding of antigen to the B-cell antigen receptor (BCR) triggers signaling that ultimately leads to B-cell activation. After BCR engagement and activation at the plasma membrane, phosphorylates PLCG2 at several sites, igniting the downstream signaling pathway through calcium mobilization, followed by activation of the protein kinase C (PKC) family members. PLCG2 phosphorylation is performed in close cooperation with the adapter protein B-cell linker protein BLNK. BTK acts as a platform to bring together a diverse array of signaling proteins and is implicated in cytokine receptor signaling pathways. Plays an important role in the function of immune cells of innate as well as adaptive immunity, as a component of the Toll-like receptors (TLR) pathway. The TLR pathway acts as a primary surveillance system for the detection of pathogens and are crucial to the activation of host defense. Especially, is a critical molecule in regulating TLR9 activation in splenic B-cells. Within the TLR pathway, induces tyrosine phosphorylation of TIRAP which leads to TIRAP degradation. BTK also plays a critical role in transcription regulation. Induces the activity of NF-kappa-B, which is involved in regulating the expression of hundreds of genes. BTK is involved on the signaling pathway linking TLR8 and TLR9 to NF-kappa-B. Transiently phosphorylates transcription factor GTF2I on tyrosine residues in response to BCR. GTF2I then translocates to the nucleus to bind regulatory enhancer elements to modulate gene expression. ARID3A and NFAT are other transcriptional target of BTK. BTK is required for the formation of functional ARID3A DNA-binding complexes. There is however no evidence that BTK itself binds directly to DNA. BTK has a dual role in the regulation of apoptosis.

GENE ID 695
SWISS PROT Q06187
SYNONYMS AGMX1; AT; ATK; BPK; IMD1; PSCTK1; XLA; Bruton Agammaglobulinemia Tyrosine Kinase; B-cell progenitor kinase


Materials Supplied

Kit Components 96 Wells Quantity/Size
Pre-coated, ready-to-use 96-well strip plate 1 plate
Plate sealer for 96 wells 2
Standard
2 tubes
Diluent buffer 1 bottle
Detection Reagent A 1 bottle
Detection Reagent B 1 bottle
TMB Substrate 1 tube
Stop Solution 1 tube
Wash Buffer (30 ℅ concentrate) 1 tube
Product data sheet 1 copy

Storage

Storage The TMB Substrate, Wash Buffer (30X concentrate), and the Stop Solution should be stored at 4°C upon receipt, while the other items should be stored at -20°C.

Performance Characteristics

REPEATABILITY

Intra-assay Precision (Precision within an assay): 3 samples with low, middle, and high-level Btk were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, middle, and high-level Btk were tested on 3 different plates, with 8 replicates in each plate.
CV(%) = SD/meanX100

Intra-Assay: CV<10%
Inter-Assay: CV<12%

SENSITIVITY The minimum detectable dose was 0.059ng/mL.
ASSAY RANGE 0.156-10ng/mL
SPECIFICITY This assay has high sensitivity and excellent specificity for the detection of Btk.
No significant cross-reactivity or interference between Btk and analogs was observed.
Note:
Limited by current skills and knowledge, it is impossible to perform all possible cross-reactivity detection tests between Btk and all analogs, therefore, cross reactivity may still exist.