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Caspase-3 Activation - An Indicator of Apoptosis in Image-Based Assays

Introduction

The understanding of the cellular signaling processes leading to programmed cell death (apoptosis) is of utmost importance in the study of autoimmune diseases such as rheumatoid arthritis or Parkinsons and Alzheimers disease. Here, errors in the signaling cascade are believed to lead to premature apoptosis in the affected tissue. At the same time, damaged cells that do not enter the apoptotic pathway may proliferate unchecked and become a cancerous cell mass. Thus, the study of apoptosis benefits many different fields of medicine by offering multiple targets for specific drug treatment.

Caspases (cysteinyl aspartate-specific proteases) are a family of important signaling molecules with various tasks depending on subtype and organ. The activation of caspases is also a marker for cellular damage in diseases such as stroke or myocardial infarction (reviewed by Lavrik et al . 2005). Although the precise role in the initiation and progression of apoptosis is not known for all caspases, their involvement as an indicator alone and as a potential leverage point for drug treatment make them widely researched molecules.

We explored the use of the active form of caspase-3 for the detection of apoptotic events. This protease has been implicated as an effector caspase associated with the initiation of the death cascade and is therefore an important marker of the cells entry point into the apoptotic signaling pathway (Nicholson et al . 1995). Caspase-3 is activated by the upstream caspase-8 and caspase-9 and since it serves as a convergence point for different signaling pathways, it is well suited as a read-out in an apoptosis assay. Detection of active caspase-3 can be used in different cell lines or primary cells, does not require the use of transfection techniques and can be multiplexed with other probes to get an in-depth understanding of signaling events with cell-by-cel
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