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Applications: Screen for molecules that inhibit TET2 activity.
Background: TET2 (also known as Tet Methylcytosine dioxygenase 2) belongs to the Ten Eleven Translocation (TET) family of proteins that catalyze 5-methylcytosine oxidation and generate 5-methylcytosine derivatives, including 5-hydroxymethylcytosine. TET2 regulates gene expression levels in cancer and immune cells, being a widely recognized tumor suppressor gene. Both homozygous and heterozygous mutations in TET2 are indicative of hematopoietic malignancies, with nonsense and frameshift mutations associated with poor prognosis. In addition, TET2 is one of the most frequently mutated genes in acute myeloid leukemia (AML). Targeting TET2 as therapeutical approach may be complex, as in many cancer types TET2 is inactivated, however it can still be found in some immune cell types and it can be modulated via proteins that regulate TET2 activity.
Contraindications: The TET2 Homogenous Assay Kit is compatible with up to 1% final DMSO concentration. We recommend preparing the inhibitor in no higher than 4% DMSO solution in buffer and using 2.5 µl per well. Green and blue dyes that absorb light in the AlphaScreen® signal emission range (520-620 nm), such as Trypan Blue, interfere with the assay. Avoid using potent singlet oxygen quenchers such as sodium azide (NaN3) or metal ions (Fe2+, Fe3+, Cu2+, Zn2+ and Ni2+). The presence of the culture medium RPMI 1640 at >1% leads to signal reduction due to the presence of excess biotin and iron in this medium. MEM, which lacks these components, does not affect AlphaScreen® assays.
Description: The TET2 (Tet Methylcytosine dioxygenase 2) Homogeneous Assay Kit is designed to measure the activity of TET2 for screening and profiling applications. The TET2 Homogeneous Assay Kit comes in a convenient AlphaLISA® format, with enough biotinylated TET2 substrate, TET2 assay buffer, and purified TET2 for 384 reactions.The key to the TET2 Homogeneous Assay Kit is a highly specific antibody that recognizes the hydroxymethylated substrate, and the increase in Alpha-counts is proportional to the hydroxymethylation of the substrate. First, a sample containing TET2 enzyme is incubated with the reaction mixture. Next, acceptor beads are added, followed by donor beads and reading the Alpha-counts.
Storage Stability: This assay kit will perform optimally for up to 6 months from date of receipt when the materials are stored as directed.
Warnings: Avoid freeze/thaw cycles
Biosafety Level: Not applicable (BSL-1)
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