This Article Full Text (PDF) References Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Add to Saved Citations Download to citation manager Request Permissions Request Reprints Add to My Marked Citations Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Citing Articles via Scopus Google Scholar Articles by Fennell, M. Articles by Wood, A. Search for Related Content PubMed PubMed Citation Articles by Fennell, M. Articles by Wood, A. Social Bookmarking                         What's this?

Multiparameter Measurement of Caspase 3 Activation and Apoptotic Cell Death in NT2 Neuronal Precursor Cells Using High-Content Analysis

Wyeth Research CN8000 Princeton, NJ 08543 Fennelm{at}wyeth.com

Helen Chan

Andrew Wood

Caspase activation is a component of a number of neurodegenerative disorders, including stroke. In this study, the authors describe a multiplexed assay for caspase 3 activation, nuclear condensation, and cell viability in a neuronal precursor cell line Ntera-2, injuredwith staurosporine and etoposide. Using a high-content screening approach, cells were identified by staining with the nuclear stain Hoechst 33342; cell viability wasmeasured by staining cells with YoPro-1, which is taken up by damaged cells but excluded from healthy cells; and caspase 3/7 activation was detected using the cell-permeable probe PhiPhi-Lux, which becomes fluorescentwhen cleaved by active caspase 3 or 7. These 3 dyeswere detected simultaneously using a 4-band pass filter set on a Cellomics Arrayscan. The authors used peptide-fmk inhibitors selective for a variety of caspases, demonstrating that the injury is mediated primarily through caspase 3 or 7, although other caspases or related proteases may play aminor role. The general caspase inhibitor zVAD-fmkwas able to block cell death and caspase activationwith the highest potency. The caspase 3 selective inhibitor DEVD-fmkwas almost as potent as zVAD-fmk; other peptide caspase inhibitors displayed onlymodest inhibition of cell death. This assay was also used as a high-content screening tool for the evaluation of novel caspase 3 inhibitors for the potential treatment of degenerative disorders.

Key Words: caspase • high-content screening • Ntera-2 • fluorescence

Journal of Biomolecular Screening, Vol. 11, No. 3, 296-302 (2006)
DOI: 10.1177/1087057105284618


CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    Twitter    What's this?

This article has been cited by other articles:

				C. Antczak, T. Takagi, C. N. Ramirez, C. Radu, and H. Djaballah Live-Cell Imaging of Caspase Activation for High-Content Screening J Biomol Screen, September 1, 2009; 14(8): 956 - 969. [Abstract] [PDF]