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Proteasome-Glo™ Chymotrypsin-Like, Trypsin-Like and Caspase-Like Cell-Based...

Proteasome-Glo™ Chymotrypsin-Like, Trypsin-Like and Caspase-Like Cell-Based Assays Technical Bulletin

Literature # TB346

The Proteasome-Glo™ Cell-Based Assay is a homogeneous, luminescent assay that measures the chymotrypsin-like, trypsin-like and caspase-like activities associated with the proteasome complex in cultured cells. The Proteasome-Glo™ Cell-Based Reagents each contain a specific luminogenic proteasome substrate in a buffer optimized for cell permeabilization, proteasome activity and luciferase activity. These peptide substrates are Suc-LLVY-aminoluciferin (Succinyl-leucine-leucine-valine-tyrosine-aminoluciferin), Z-LRR-aminoluciferin (Z-leucine-arginine-arginine-aminoluciferin) and Z-nLPnLD-aminoluciferin (Z-norleucine-proline-norleucine-aspartate-aminoluciferin) for the chymotrypsin-like, trypsin-like and caspase-like activities, respectively. The trypsin-like assay also contains two inhibitors to reduce nonspecific protease activities. Adding a single Proteasome-Glo™ Cell-Based Reagent in an “add-mix-measure” format results in proteasome cleavage of the substrate and generation of a luminescent signal by the luciferase reaction. The Proteasome-Glo™ Reagent contains the proprietary thermostable luciferase, Ultra-Glo™ Recombinant Luciferase, and is formulated to generate a stable, “glow-type” luminescent signal that improves performance across a wide range of assay conditions. This coupled enzyme system, with simultaneous proteasome cleavage of substrate and luciferase consumption of the released aminoluciferin, results in a luminescent signal that is proportional to the amount of proteasome activity in cells. Steady state activities of the proteasome and luciferase enzymes are reached within 5–10 minutes after adding the reagent, allowing fast and easy monitoring of activity.

Printed in USA. Revised 12/12.

 

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