Apoptosis-associated caspase activation assays

Scott H. Kaufmann, Sun Hee Lee, X. Wei Meng, David A. Loegering, Timothy J. Kottke, Alexander J. Henzing, Sandrine Ruchaud, Kumiko Samejima, William C. Earnshaw

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62 Scopus citations

Abstract

Caspases are aspartate-directed cysteine proteases that cleave a diverse group of intracellular substrates to contribute to various manifestations of apoptosis. These proteases are synthesized as inactive precursors and are activated as a consequence of signaling induced by a wide range of physiological and pathological stimuli. Caspase activation can be detected by measurement of catalytic activity, immunoblotting for cleavage of their substrates, immunolabeling using conformation-sensitive antibodies or affinity labeling followed by flow cytometry or ligand blotting. Here we describe methods for each of these assays, identify recent improvements in these assays and outline the strengths and limitations of each approach.

Original languageEnglish (US)
Pages (from-to)262-272
Number of pages11
JournalMethods
Volume44
Issue number3
DOIs
StatePublished - Mar 1 2008

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Keywords

  • Affinity labeling
  • Apoptosis
  • Caspases
  • Enzymatic activity
  • Flow cytometry
  • Fluorogenic substrate
  • Immunoblotting
  • Immunofluorescence

ASJC Scopus subject areas

  • Molecular Biology
  • Biochemistry, Genetics and Molecular Biology(all)

Cite this

Kaufmann, S. H., Lee, S. H., Meng, X. W., Loegering, D. A., Kottke, T. J., Henzing, A. J., Ruchaud, S., Samejima, K., & Earnshaw, W. C. (2008). Apoptosis-associated caspase activation assays. Methods, 44(3), 262-272. https://doi.org/10.1016/j.ymeth.2007.11.005