Caspases are amongst the most specific endopeptidases known today. Therefore, activation of this family of proteases results in limited proteolytic cleavage at specific aspartate residues of the target molecules (33). A novel approach to probe for specific proteolytic events during apoptosis is the use of antibodies directed against neo-epitopes generated by caspase cleavage. This approach has been shown to be feasible for the caspase-cleaved cytoskeletal proteins actin and cytokeratin 18. Yang et al. (14) raised a polyclonal antiserum using a synthetic peptide representing the last five amino acids of the C-terminus of the 32 kDa actin fragment produced during apoptosis. The antiserum specifically labels apoptotic but not necrotic cells. Similarly, Leers et al. (25) describe a monoclonal antibody specific for the liberated DALD sequence in the C-terminus of cytokeratin 18. Also this antibody specifically recognizes apoptotic cells and does not detect necrotic cells. Antibodies for the in situ detection of caspase-cleaved cyto- and nucleoskeletal proteins provide simple, specific tools to evaluate the role of caspase activation in cytoskeletal reorganization during apoptosis. They are now in use as apoptosis markers that can be applied to routinely processed tissues and cells.
3. Apoptosis detection systems
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