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ITK Kinase Assay

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Compound Test Services

CT-001

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Product Description


ITK (Interleukin-2-inducible T-cell kinase), also known as EMT or TSK, is a member of the non-receptor tyrosine kinase family (Tec kinase subfamily) that plays a pivotal role in T-cell receptor (TCR) signaling, Th2 differentiation, and immune responses. ITK is activated by various stimuli including TCR engagement and co-stimulatory signals, leading to phosphorylation of PLCγ1, calcium influx, and activation of NF-AT, NF-κB, and MAPK pathways, thereby promoting T-cell activation, proliferation, and cytokine production. Beyond its role in normal adaptive immunity, dysregulation of ITK signaling—through loss-of-function mutations or overexpression—is closely associated with primary immunodeficiency (e.g., X-linked lymphoproliferative disease), allergic asthma, and multiple cancers (including T-cell leukemia and lymphoma), making it a highly validated target for immunosuppressive therapy, asthma treatment, and cancer drug discovery.


Screeningbio’s ITK Kinase Assay Kit provides a robust, sensitive, and high-throughput platform for quantifying ITK kinase activity and evaluating the potency of tyrosine kinase inhibitors (TKIs).


This kit utilizes the ADP-Glo Kinase Assay platform, a luminescent technology that measures the amount of ADP produced during the kinase reaction.

The assay is performed in two distinct steps:

  1. Kinase Reaction: ITK catalyzes the transfer of a phosphate group from ATP to a specific substrate, resulting in the production of phosphorylated substrate and ADP.

  2. ADP-Glo™ Reagent Addition: After the kinase reaction, the ADP-Glo™ Reagent is added to terminate the reaction and deplete any remaining unreacted ATP.

  3. Kinase Detection: The Kinase Detection Reagent is added to simultaneously convert the produced ADP back into ATP, which is then used by luciferase to generate a luminescent signal.

The light intensity produced is directly proportional to the ADP concentration and, consequently, to the enzymatic activity of the ITK  kinase.




Data

TKs Kinase Inhibition Assay. Staurosporine was titrated using established assay protocol. Non-linear regression was used to plot RLU signal vs. [Compound, M], and EC50 /IC50 values were determined, using GraphPad Prism software.
TKs Kinase Inhibition Assay. Staurosporine was titrated using established assay protocol. Non-linear regression was used to plot RLU signal vs. [Compound, M], and EC50 /IC50 values were determined, using GraphPad Prism software.


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