JMJD3 (KDM6B) Homogeneous Assay Kit

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Catalog #

50416

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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.

Scheme 1

Need us to run inhibitor screens or profile your compounds against JMJD3 (KDM6B)? Check out our Demethylase Screening Services.

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Synonyms

JMJD3, KDM6B

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Product Data Gallery

Product Info

Storage and Usage

Citations

Assay Kit Format

AlphaLISA®

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.

Materials Required But Not Supplied

AlphaLISA® anti-rIgG acceptor beads, 5 mg/ml (PerkinElmer #AL104C)
AlphaScreen® Streptavidin-conjugated donor beads, 5 mg/ml (PerkinElmer #6760002S)
Optiplate -96 (PerkinElmer #6005290) or Optiplate -384 (PerkinElmer #6007290)
AlphaScreen® microplate reader
Adjustable micropipettor and sterile tips

Format

96 Reactions:

Catalog Number	Component	Amount	Storage
50115	JMJD3 (KDM6B)	4 µg	-80°C	Avoid *multiple* *freeze/thaw* *cycles!*
52140F	Primary antibody 6	5 µl	-80°C
79841	Biotinylated histone H3 peptide substrate	300 rxns	-80°C
52408	4x HDM Assay Buffer 3	3 ml	-80°C
52301	4x Detection buffer	2 ml	-20°C

384 Reactions:

Catalog Number	Component	Amount	Storage
50115	JMJD3 (KDM6B)	20 µg	-80°C	Avoid *multiple* *freeze/thaw* *cycles!*
52140F	Primary antibody 6	20 µl	-80°C
79841	Biotinylated histone H3 peptide substrate	500 rxns	-80°C
52408	4x HDM Assay Buffer 3	3 ml	-80°C
52301	4x Detection buffer	2 ml	-20°C

UniProt #

O15054

Background

JMJD3 is a JmjC-domain protein that exhibits demethylation activity toward di- and trimethyl-lysine 27 (H3K27me2/3) on histone H3.

References

Swigut T., Wysocka J. Cell 2007; 131(1): 29-32.

Storage/Stability

6 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.

Instructions for Use

See assay kit data sheet for detailed protocol.

Applications

Useful for studying enzyme kinetics and screening small molecular inhibitors for drug discovery and HTS applications.

Shipping Temperature

-80°C

Notes

AlphaScreen^® and AlphaLISA^® are registered trademarks of PerkinElmer, Inc.

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.

Warnings

Avoid freeze/thaw cycles.

Disclaimers and Limitations

BPS Bioscience assay kits are validated using the listed components according to our specific protocol. Any deviations to kit components or protocols, such as using different proteins, cell/tissue lysates, or buffers (including using the same component from other commercial sources) will void any warranties on the performance of the assay kit and is not recommended.

End-users should immediately report any issues to [email protected] upon receipt of the kit, such as missing components, damaged vials, no dry ice, etc.

Scientific Category

Demethylase

Regulatory Status

Research Use Only

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JMJD3 (KDM6B) Homogeneous Assay Kit

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