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A quantitative method for the specific assessment of caspase-6 activity in cell culture

Publikation: Bidrag til tidsskriftTidsskriftartikelForskningfagfællebedømt

  • Dagmar E Ehrnhoefer
  • Skotte, Niels
  • Jane Savill
  • Yen T N Nguyen
  • Safia Ladha
  • Li-Ping Cao
  • Edie Dullaghan
  • Michael R Hayden

Aberrant activation of caspase-6 has recently emerged as a major contributor to the pathogeneses of neurodegenerative disorders such as Alzheimer's and Huntington disease. Commercially available assays to measure caspase-6 activity commonly use the VEID peptide as a substrate. However these methods are not well suited to specifically assess caspase-6 activity in the presence of other, confounding protease activities, as often encountered in cell and tissue samples. Here we report the development of a method that overcomes this limitation by using a protein substrate, lamin A, which is highly specific for caspase-6 cleavage at amino acid 230. Using a neo-epitope antibody against cleaved lamin A, we developed an electrochemiluminescence-based ELISA assay that is suitable to specifically detect and quantify caspase-6 activity in highly apoptotic cell extracts. The method is more sensitive than VEID-based assays and can be adapted to a high-content imaging platform for high-throughput screening. This method should be useful to screen for and characterize caspase-6 inhibitor compounds and other interventions to decrease intracellular caspase-6 activity for applications in neurodegenerative disorders.

OriginalsprogEngelsk
TidsskriftPLOS ONE
Vol/bind6
Udgave nummer11
Sider (fra-til)e27680
ISSN1932-6203
DOI
StatusUdgivet - 2011

ID: 153451326