A quantitative method for the specific assessment of caspase-6 activity in cell culture.
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 met...
| Main Authors: | , , , , , , , |
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| Format: | Article |
| Language: | English |
| Published: |
Public Library of Science (PLoS)
2011-01-01
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| Series: | PLoS ONE |
| Online Access: | http://europepmc.org/articles/PMC3226564?pdf=render |
| _version_ | 1818057083008843776 |
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| author | Dagmar E Ehrnhoefer Niels H Skotte Jane Savill Yen T N Nguyen Safia Ladha Li-Ping Cao Edie Dullaghan Michael R Hayden |
| author_facet | Dagmar E Ehrnhoefer Niels H Skotte Jane Savill Yen T N Nguyen Safia Ladha Li-Ping Cao Edie Dullaghan Michael R Hayden |
| author_sort | Dagmar E Ehrnhoefer |
| collection | DOAJ |
| description | 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. |
| first_indexed | 2024-12-10T12:39:05Z |
| format | Article |
| id | doaj.art-a38f098eb9794746a35f3a623c977d86 |
| institution | Directory Open Access Journal |
| issn | 1932-6203 |
| language | English |
| last_indexed | 2024-12-10T12:39:05Z |
| publishDate | 2011-01-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS ONE |
| spelling | doaj.art-a38f098eb9794746a35f3a623c977d862022-12-22T01:48:35ZengPublic Library of Science (PLoS)PLoS ONE1932-62032011-01-01611e2768010.1371/journal.pone.0027680A quantitative method for the specific assessment of caspase-6 activity in cell culture.Dagmar E EhrnhoeferNiels H SkotteJane SavillYen T N NguyenSafia LadhaLi-Ping CaoEdie DullaghanMichael R HaydenAberrant 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.http://europepmc.org/articles/PMC3226564?pdf=render |
| spellingShingle | Dagmar E Ehrnhoefer Niels H Skotte Jane Savill Yen T N Nguyen Safia Ladha Li-Ping Cao Edie Dullaghan Michael R Hayden A quantitative method for the specific assessment of caspase-6 activity in cell culture. PLoS ONE |
| title | A quantitative method for the specific assessment of caspase-6 activity in cell culture. |
| title_full | A quantitative method for the specific assessment of caspase-6 activity in cell culture. |
| title_fullStr | A quantitative method for the specific assessment of caspase-6 activity in cell culture. |
| title_full_unstemmed | A quantitative method for the specific assessment of caspase-6 activity in cell culture. |
| title_short | A quantitative method for the specific assessment of caspase-6 activity in cell culture. |
| title_sort | quantitative method for the specific assessment of caspase 6 activity in cell culture |
| url | http://europepmc.org/articles/PMC3226564?pdf=render |
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