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Applications: Useful for studying enzyme kinetics and screening small molecular inhibitors for drug discovery and HTS applications.
Background: JARID1A, also known as RBBP2 and KDM5A, is a JumonjiC (JmjC) and ARID domain-containing histone lysine demethylase that exhibits demethylation activity toward di- and trimethyl-lysine 4 (H3K4me2/3) on histone H3.
Contraindications: Avoid green and blue dyes that absorb light in the Alphascreen® signal emission range (520-620 nm), such as Trypan Blue. Avoid the use of the potent singlet oxygen quenchers such as sodium azide (NaN3) or metal ions (Fe2+, Fe3+, Cu2+, Zn2+ and Ni2+). The presence of >1% RPMI 1640 culture medium leads to a 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 JARID1A Homogeneous Assay Kit is designed to measure JARID1A activity for screening and profiling applications. JARID1A, also known as RBBP2 and KDM5A, is a JumonjiC (JmjC) and ARID domain-containing histone lysine demethylase that exhibits demethylation activity toward H3-K4Me2 and H3-K4Me3. The JARID1A Homogeneous Assay Kit comes in a convenient AlphaLISA® format (Scheme 1), with biotinylated histone H3 peptide substrate, primary antibody, demethylase assay buffer, and purified JARID1A for 96 or 384 enzyme reactions. The key to the JARID1A Homogeneous Assay Kit is a highly specific antibody that recognizes demethylated substrate. With this kit, only three simple steps on a microtiter plate are required for methyltransferase detection. First, a sample containing JARID1A enzyme is incubated with the biotinylated substrate. Next, acceptor beads and primary antibody are added, then donor beads, followed by reading the Alpha-counts. Our histone demethylase assays utilize highly specific antibodies that recognize demethylated histone substrate. First, a sample containing the enzyme is incubated with a biotinylated substrate. Next, acceptor beads and primary antibody are added, then donor beads, followed by reading the Alpha-counts.
Storage Stability: At least 6 months at -80°C.
Supplied As: The JARID1A Homogeneous Assay Kit comes in a convenient AlphaLISA® format, with biotinylated histone H3 peptide substrate, primary antibody, demethylase assay buffer, and purified JARID1A for 96 or 384 enzyme reactions.
Warnings: Avoid freeze/thaw cycles.
Biosafety Level: Not applicable (BSL-1)
References: 1. DiTacchio L, Le HD, Vollmers C et al. Science 2011; 333(6051):1881-5.
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