The BRG1 Inhibitor Screening Assay Kit is designed to measure the inhibition of BRG1 binding to its substrate. The key to the BRG1 Inhibitor Screening Assay Kit is the specific binding of the BRG1 bromodomain to an acetylated histone substrate. With this kit, only three simple steps on a microtiter plate are required. First, a sample containing BRG1 and an inhibitor of choice is incubated with the biotinylated substrate. Next, acceptor beads are added, then donor beads, followed by reading the Alpha-counts.

MATERIALS OR INSTRUMENTS REQUIRED BUT NOT SUPPLIED:
Glutathione AlphaLISA^® Acceptor Beads, 5 mg/ml (PerkinElmer #AL109C)
AlphaScreen^® Streptavidin-conjugated donor beads, 5 mg/ml (PerkinElmer #6760002)
Optiplate-384 (PerkinElmer #6007290)
AlphaScreen^® microplate reader
Adjustable micropipettor and sterile tips

BRG1, SMARCA4, activity assay kit

The kit comes in a convenient AlphaLISA^® format, with biotinylated histone peptide substrate, assay buffer, detection buffer and purified GST-tagged BRG1 bromodomain to perform a total of 384 enzyme reactions.

DMSO above 0.5%. Only limited amounts of DMSO can be included, as it has been shown to disrupt BRD-ligand interaction. 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.

COMPONENTS

See assay kit data sheet for detailed protocol.

At least 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.

Useful for the study of bromodomain binding assays, screening bromodomain inhibitors and selectivity profiling.

Filippakopoulos, P., et al., Cell 2012; 149:214.

MATERIALS OR INSTRUMENTS REQUIRED BUT NOT SUPPLIED:
Glutathione AlphaLISA^® Acceptor Beads, 5 mg/ml (PerkinElmer #AL109C)
AlphaScreen^® Streptavidin-conjugated donor beads, 5 mg/ml (PerkinElmer #6760002)
Optiplate-384 (PerkinElmer #6007290)
AlphaScreen^® microplate reader
Adjustable micropipettor and sterile tips

Avoid freeze/thaw cycles.

Bromodomain

Assay Kit