A high-throughput direct fluorescence resonance energy transfer-based assay for analyzing apoptotic proteases using flow cytometry and fluorescence lifetime measurements

Analytical Biochemistry
Miho SuzukiMaïté Coppey-Moisan

Abstract

Cytometry is a versatile and powerful method applicable to different fields, particularly pharmacology and biomedical studies. Based on the data obtained, cytometric studies are classified into high-throughput (HTP) or high-content screening (HCS) groups. However, assays combining the advantages of both are required to facilitate research. In this study, we developed a high-throughput system to profile cellular populations in terms of time- or dose-dependent responses to apoptotic stimulations because apoptotic inducers are potent anticancer drugs. We previously established assay systems involving protease to monitor live cells for apoptosis using tunable fluorescence resonance energy transfer (FRET)-based bioprobes. These assays can be used for microscopic analyses or fluorescence-activated cell sorting. In this study, we developed FRET-based bioprobes to detect the activity of the apoptotic markers caspase-3 and caspase-9 via changes in bioprobe fluorescence lifetimes using a flow cytometer for direct estimation of FRET efficiencies. Different patterns of changes in the fluorescence lifetimes of these markers during apoptosis were observed, indicating a relationship between discrete steps in the apoptosis process. The finding...Continue Reading

References

May 2, 1994·Journal of Immunological Methods·A B Lyons, C R Parish
May 28, 1996·Proceedings of the National Academy of Sciences of the United States of America·J T PaiJ L Goldstein
Aug 10, 2004·Biophysical Journal·Marisa Martin-FernandezGareth R Jones
Feb 22, 2005·Current Opinion in Biotechnology·Horst Wallrabe, Ammasi Periasamy
Mar 23, 2005·Nature Methods·Emilia GalperinAlexander Sorkin
Jul 19, 2005·Journal of Immunological Methods·Kristina E HowardMary Jo Burkhard
Sep 16, 2005·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·Peter NagyJános Szöllősi
Sep 22, 2005·Cancer Chemotherapy and Pharmacology·Ryungsa KimKazuaki Tanabe
May 10, 2006·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·Xiaoli WuLiusheng He
May 29, 2007·FEBS Letters·Hyung-Seok ChoiKwang-Hyun Cho
Aug 21, 2009·European Biophysics Journal : EBJ·A J W G VisserJ W Borst
Oct 22, 2009·Biophysical Journal·Sergi Padilla-ParraMarc Tramier
Jun 22, 2010·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·Johannes T WesselsFred S Wouters
Sep 23, 2010·Laboratory Hematology : Official Publication of the International Society for Laboratory Hematology·Howard J Meyerson
Feb 22, 2012·Journal of Visualized Experiments : JoVE·Klaus SuhlingGokhan Yahioglu
Apr 17, 2012·PloS One·Daniel R MatthewsSimon M Ameer-Beg
Jul 17, 2012·Journal of Laboratory Automation·Constantin RaduHakim Djaballah
Apr 16, 2013·Methods : a Companion to Methods in Enzymology·Giulia Freer, Laura Rindi
Nov 30, 2013·TheScientificWorldJournal·Azhar RasulXiaomeng Li
Feb 25, 2014·Tumour Biology : the Journal of the International Society for Oncodevelopmental Biology and Medicine·Jun LiuRenan Chen
Mar 19, 2014·Advanced Drug Delivery Reviews·Claudio R ThomaWilhelm Krek
Mar 19, 2014·International Journal of Cell Biology·Stefan W RyterAugustine M K Choi
May 17, 2014·Assay and Drug Development Technologies·Rasheena EdmondsonLiju Yang
May 31, 2014·Cell Death & Disease·I N Lavrik
Jul 16, 2014·Proceedings of the National Academy of Sciences of the United States of America·Mathias J WawerPaul A Clemons
Aug 12, 2014·The Journal of Biological Chemistry·Maximilian L Würstle, Markus Rehm
Aug 19, 2014·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·Michael HalterJohn T Elliott

❮ Previous
Next ❯

Citations

Nov 24, 2016·SLAS Discovery·Tory M SchaafGregory D Gillispie
Jul 1, 2017·Methods and Applications in Fluorescence·Franz-Josef Meyer-Almes
Aug 14, 2020·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·Kapil NichaniJessica P Houston

❮ Previous
Next ❯

Related Concepts

Related Feeds

Apoptotic Caspases

Apoptotic caspases belong to the protease enzyme family and are known to play an essential role in inflammation and programmed cell death. Here is the latest research.

Apoptosis

Apoptosis is a specific process that leads to programmed cell death through the activation of an evolutionary conserved intracellular pathway leading to pathognomic cellular changes distinct from cellular necrosis