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AlphaLISA technology is a powerful and versatile platform that offers, highly sensitive, no-wash immunoassays using Alpha Donor and, AlphaLISA Acceptor beads. In this technical note, we present the, optimization of an HDAC1 enzymatic assay using a biotinylated, histone H3K9ac peptide as substrate. Detection of the deacetylated, product was performed by the addition of Streptavidin (SA) Alpha, Donor beads and AlphaLISA Acceptor beads conjugated to an antibody, (Ab) directed against the unmodified H3K9 residue. Upon laser, irradiation of the beads-target complexes at 680 nm, short-lived, singlet oxygen molecules produced by the Donor beads can reach the, Acceptor beads in proximity to generate an amplified chemiluminescent, signal at 615 nm. The intensity of the light emission is proportional to, the deacetylation activity of the HDAC1 enzyme.
This AlphaLISA immunodetection assay measures the demethylation, of a biotinylated histone H3 (1-21) peptide mono-methylated at, lysine 9.
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