2012 Promega catalogue

Cell Signaling
Cell-Based Proteasome-Glo Assays
Product Size Cat.# Price ($)
Proteasome-Glo Chymotrypsin-Like Cell-
10 ml G8660 751.00
Based Assay
5 × 10 ml G8661 Pls. Enq.
2 × 50 ml G8662 Pls. Enq.
Proteasome-Glo Trypsin-Like Cell-Based
10 ml G8760 751.00
Assay
5 × 10 ml G8761 Pls. Enq.
Proteasome-Glo Caspase-Like Cell-Based 10 ml G8860 751.00
Assay
5 × 10 ml G8861 Pls. Enq.
Proteasome-Glo 3-Substrate Cell-Based Assay 10 ml G1180 1741.00
System
For Laboratory Use.
50 ml G1200 Pls. Enq.
Description: The Proteasome-Glo Cell-Based Assays are homogeneous,
luminescent assays that individually measure the chymotrypsin-like, trypsin-like
and caspase-like protease activities associated with the proteasome complex in
cultured cells. The 26S proteasome is a 2.5MDa multiprotein complex found in
all eukaryotic cells. Proteasome-Glo Cell-Based Assays provide luminogenic
proteasome substrates in buffers optimized for cell permeabilization,
proteasome activity and luciferase activity. Addition of the Proteasome-Glo
Cell-Based Reagent in an “add-mix-measure” format results in proteasome
cleavage of the substrate and rapid generation of a luminescent signal
produced by the luciferase reaction.
The three luminogenic substrates used to monitor specific protease activities
include: Suc-LLVY-aminoluciferin for chymotrypsin-like, Z-LRR-aminoluciferin
for trypsin-like, and Z-nLPnLD-aminoluciferin for caspase-like activity. Each
luminogenic substrate is added to a buffer system optimized for its specific
proteasome activity and luciferase activity. The reagents are added to cells in
culture, and the proteasome cleaves the substrates, releasing luciferin, which
is consumed by luciferase, producing “glow-type” luminescence correlating to
enzyme activity or inhibition.
The Proteasome-Glo Cell-Based 3-Substrate System consists of three
homogeneous bioluminescent assays that measure the three proteolytic
activities associated with the proteasome in a cell-based format (each of these
three assays also is available separately).
The Proteasome-Glo 3-Substrate System consists of three
homogeneous bioluminescent assays in an enzyme-based format (each of
these three assays also is available separately).
Features:
• More Biologically Relevant Results: Obtain activity data directly from a
cellular environment with the Proteasome-Glo Cell-Based Assay.
• Simplified Method: The “add-mix-measure” protocol minimizes handling
steps and makes the assays amenable to automation.
• Faster Results: Maximum sensitivity is reached 10–30 minutes after
reagent addition.
• Greater Sensitivity: The luminescent assay format avoids inherent
fluorescent background signals, providing excellent signal-to-background
readings. The assays are miniaturizable to 384-well format.
Storage Conditions: Store the Proteasome-Glo Assay components at
–20°C.
Protocol Part#
Proteasome-Glo Chymotrypsin-Like, Trypsin-Like and Caspase-
Like Cell-Based Assays Technical Bulletin
TB346
Proteasome
For complete and up-to-date product information visit: www.promega.com/catalog
H
Suc-LLVY – N
or
Z-LRR –
or
Z-nLPnLD –
S
N
Suc-LLVY + H2N or
S N COOH
Z-LRR
or
Z-nLPnLD
N S
Aminoluciferin
Ultra-Glo rLuciferase
Light
N
S
COOH
ATP, Mg ++ , O 2
The luminogenic substrates containing the Suc-LLVY, Z-LRR or
ZnLPnLD sequence are recognized by the 20S proteasome. Following
cleavage by the 20S proteasome, the substrate for luciferase (aminoluciferin)
is released, allowing the luciferase reaction to produce light.
Signal-to-Noise Ratio
100,000
10,000
1,000
100
10
Suc-LLVY-Glo Substrate
Z-LRR-Glo Substrate
Z-nLPnLD-Glo Substrate
Suc-LLVY-AMC
Suc-LLVY-AMC+SDS
Boc-LRR-AMC
Ac-nLPnLD-AMC
1
0.00001 0.0001 0.001 0.01 0.1
20S (µg/ml)
Luminescent proteasome assays are more sensitive than fluorescent
proteasome assays.
1
10
5855MA
5994MB
143
6
Drug Discovery
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