2013 Promega catalogue

Cell Signaling
SIRT-Glo Assays and Screening Systems
Product Size Cat.# Price ($)
SIRT-Glo Assay 10 ml G6450 584
5 × 10 ml G6451 2159
100 ml G6452 2809
SIRT-Glo Screening System 10 ml G6470 622
5 × 10 ml G6471 2303
Available Separately Size Conc. Cat.# Price ($)
Nicotinamide 30 μl 1 M G6540 114
HeLa Nuclear Extract 10 μl 5 mg/ml G6570 167
SIRT-Glo Control Substrate 35 μl G6460 356
For Research Use Only. Not for Use in Diagnostic Procedures.
Description: The SIRT-Glo Assays and Screening Systems are singlereagent-addition,
homogeneous, luminescent assays that measure the relative
activity of the NAD + -dependent histone deacetylase (HDAC) class III enzymes
(sirtuins; SIRTs) from purified enzyme sources. The assays use an acetylated,
luminogenic peptide substrate that can be deacetylated by SIRT activities.
Deacetylation of the peptide aminoluciferin substrate is measured using a
coupled enzymatic system in which a protease in the Developer Reagent
cleaves the deacetylated peptide from the aminoluciferin substrate, releasing
aminoluciferin, which is quantified in a reaction using Ultra-Glo recombinant
firefly luciferase. The assay reaction is typically complete within 15–45
minutes with no sample manipulation. The SIRT-mediated luminescent signal is
persistent with a half-life of greater than 3 hours, allowing batch processing of
multiwell plates. The SIRT-Glo Assay is broadly useful for NAD + -dependent
Sirtuin enzymes.
Nicotinamide, included in the SIRT-Glo Screening Systems or available
separately, is a known inhibitor of SIRTs and used as a positive control inhibitor.
Nicotinamide is supplied at a concentration of 1M in SIRT-Glo Buffer.
The HeLa Nuclear Extract, included in the SIRT-Glo Screening Systems or
available separately, may be used as an assay chemistry control. HeLa Nuclear
Extract is supplied at a concentration of 5mg/ml.
The SIRT-Glo Control Substrate, only available separately, is a non-acetylated
form of the SIRT-Glo Substrate with the same amino acid sequence and can
be used with the SIRT-Glo Assays and Screening Systems to confirm true
SIRT inhibition in secondary screens. The Control Substrate is supplied at a
concentration of 10mM and is sufficient for 480 assays in 96-well plate format
when combined with the SIRT-Glo Reagent prepared with components in the
SIRT-Glo Assays or Screening Systems.
Features:
• Simple Measurement of Deacetylating Activities: Use a singlereagent-addition,
homogeneous, add-mix-measure protocol for easy
implementation from benchtop to screening.
• Highly Sensitive: Achieve 10- to 100-fold higher sensitivity than
comparable fluorescence methods.
• Fast Data Acquisition: Measure maximum signal in as little as
10-15 minutes with persistent glow-type steady-state signal.
Storage Conditions: Store the SIRT-Glo Assay components and SIRT-Glo
Control Substrate at –20°C. Store HeLa Nuclear Extract at –70°C.
Protocol
HDAC-Glo I/II Assay and Screening System Technical Manual
SIRT-Glo Assay and Screening System Technical Manual
Signal-to-Noise Ratio
10,000
1,000
100
10
SIRT1
SIRT2
SIRT3
SIRT4
SIRT5
SIRT6
1
1 10 100 1,000
Recombinant SIRT Enzyme (ng/ml)
Broad linearity and isoenzyme utility.
Part#
TM335
TM336
8910MA
4
Cell Signaling
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