Jove
Visualize
Contact Us
JoVE
x logofacebook logolinkedin logoyoutube logo
ABOUT JoVE
OverviewLeadershipBlogJoVE Help Center
AUTHORS
Publishing ProcessEditorial BoardScope & PoliciesPeer ReviewFAQSubmit
LIBRARIANS
TestimonialsSubscriptionsAccessResourcesLibrary Advisory BoardFAQ
RESEARCH
JoVE JournalMethods CollectionsJoVE Encyclopedia of ExperimentsArchive
EDUCATION
JoVE CoreJoVE BusinessJoVE Science EducationJoVE Lab ManualFaculty Resource CenterFaculty Site
Terms & Conditions of Use
Privacy Policy
Policies

Related Concept Videos

You might also read

Related Articles

Articles linked to this work by shared authors, journal, and citation graph.

Sort by
Same author

Interrogating direct NLRP3 engagement and functional inflammasome inhibition using cellular assays.

Cell chemical biology·2023
Same author

Treating Cells as Reagents to Design Reproducible Assays.

SLAS discovery : advancing life sciences R & D·2021
Same author

"Multiplexed viability, cytotoxicity, and caspase activity assays".

Methods in molecular biology (Clifton, N.J.)·2014
Same author

Update on in vitro cytotoxicity assays for drug development.

Expert opinion on drug discovery·2013
Same author

Development of a dehalogenase-based protein fusion tag capable of rapid, selective and covalent attachment to customizable ligands.

Current chemical genomics·2012
Same author

A bioluminescent assay for the sensitive detection of proteases.

BioTechniques·2011

Related Experiment Video

Updated: Apr 22, 2026

Investigating Protein-protein Interactions in Live Cells Using Bioluminescence Resonance Energy Transfer
11:46

Investigating Protein-protein Interactions in Live Cells Using Bioluminescence Resonance Energy Transfer

Published on: May 26, 2014

22.7K

Homogeneous, bioluminescent proteasome assays.

Martha A O'Brien1, Richard A Moravec, Terry L Riss

  • 1Promega Corporation, 2800 Woods Hollow Road, Madison, WI, 53711, USA, martha.obrien@promega.com.

Methods in Molecular Biology (Clifton, N.J.)
|October 14, 2014
PubMed
Summary

A new bioluminescent assay enables sensitive, single-step measurement of proteasome activity in cells. This homogeneous method is ideal for high-throughput screening of proteasome inhibitors for cancer and neurodegenerative diseases.

More Related Videos

Demonstration of Heterologous Complexes formed by Golgi-Resident Type III Membrane Proteins using Split Luciferase Complementation Assay
05:28

Demonstration of Heterologous Complexes formed by Golgi-Resident Type III Membrane Proteins using Split Luciferase Complementation Assay

Published on: September 10, 2020

1.9K
High-throughput Functional Screening using a Homemade Dual-glow Luciferase Assay
12:55

High-throughput Functional Screening using a Homemade Dual-glow Luciferase Assay

Published on: June 1, 2014

23.4K

Related Experiment Videos

Last Updated: Apr 22, 2026

Investigating Protein-protein Interactions in Live Cells Using Bioluminescence Resonance Energy Transfer
11:46

Investigating Protein-protein Interactions in Live Cells Using Bioluminescence Resonance Energy Transfer

Published on: May 26, 2014

22.7K
Demonstration of Heterologous Complexes formed by Golgi-Resident Type III Membrane Proteins using Split Luciferase Complementation Assay
05:28

Demonstration of Heterologous Complexes formed by Golgi-Resident Type III Membrane Proteins using Split Luciferase Complementation Assay

Published on: September 10, 2020

1.9K
High-throughput Functional Screening using a Homemade Dual-glow Luciferase Assay
12:55

High-throughput Functional Screening using a Homemade Dual-glow Luciferase Assay

Published on: June 1, 2014

23.4K

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Drug Discovery

Background:

  • The ubiquitin-proteasome pathway is crucial for protein degradation and cellular regulation.
  • Proteasome dysfunction is linked to cancer, neurodegenerative, and cardiovascular diseases.
  • Existing fluorogenic assays for proteasome activity have limitations, including high background and interference.

Purpose of the Study:

  • To develop a homogeneous, bioluminescent assay for monitoring proteasome activity.
  • To create a cell-based assay suitable for high-throughput screening (HTS).
  • To enable sensitive measurement of distinct proteasome catalytic activities.

Main Methods:

  • Developed a homogeneous, bioluminescent method using peptide-conjugated aminoluciferin substrates and stabilized luciferase.
  • Incorporated a selective membrane permeabilization step for cell-based assays.
  • Designed assays to measure chymotrypsin-like, trypsin-like, and caspase-like proteasome activities.

Main Results:

  • The new method provides sensitive, single-step, cell-based assays for each proteasome catalytic activity.
  • Eliminates the need for cell extract preparation, simplifying the assay workflow.
  • Demonstrated adequate sensitivity for 96- and 384-well plate formats.

Conclusions:

  • The developed bioluminescent assay is a robust and rapid tool for proteasome activity monitoring.
  • The "add and read" format is ideal for high-throughput screening of proteasome inhibitors.
  • This assay facilitates the development of therapeutics targeting proteasome-related diseases.