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Applications: Useful for studying enzyme kinetics and screening small molecular inhibitors for drug discovery and HTS applications.
Background: JMJD3 is a JmjC-domain protein that exhibits demethylation activity toward di- and trimethyl-lysine 27 (H3K27me2/3) 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 JMJD3 Homogeneous Assay Kit is designed to measure JMJD3 activity for screening and profiling applications. JMJD3, also known as KDM6B, is a JumonjiC (JmjC) domain protein that exhibits demethylation activity toward H3-K27Me3 and H3-K27Me2. The key to the JMJD3 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 JMJD3 enzyme is incubated with the biotinylated substrate. Next, acceptor beads and primary antibody are added, then donor beads, followed by reading the Alpha-counts.
Storage Stability: 12 months from date of receipt, when stored as directed. Kit components require different storage conditions. Be sure to store each component at the proper temperature upon arrival.
Supplied As: Kit comes in a convenient AlphaLISA® format, with biotinylated histone H3 peptide substrate, primary antibody, demethylase assay buffer, and purified JMJD3 for 96 or 384 enzyme reactions.
Warnings: Avoid freeze/thaw cycles.
Biosafety Level: Not applicable (BSL-1)
References: Swigut T., Wysocka J. Cell 2007; 131(1): 29-32.
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