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Title: Analysis of the role of caspase 2 in apoptosis
Author: Dost, Britta
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 1998
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In order to define the role of caspase 2 in apoptosis, rabbit polyclonal antibodies were raised against recombinant human caspase 2 fragments. Characterization of the three polyclonal caspase 2 antibodies by immunoblot and ELISA showed that all three antibodies are highly specific to recombinant caspase 2 protein. They proved, however, to have very low affinity towards endogenous human caspase 2 and were unable to detect native caspase 2 in Western blotting and immunohistochemical analysis. A commercially available anti-caspase 2 monoclonal antibody, generated against recombinant caspase 2, was shown by immunoblot analysis to detect a cellular protein of the appropriate molecular weight for procaspase 2. This antibody was used to investigate caspase 2 expression by immunoblot analysis in various human tissues and cell lines. Caspase 2 expression was found at various levels in all 11 tissues analyzed. Immunoreactivity was localized within the cytosol in several human tissues: the renal tubular epithelium, the pituitary gland, splenic lymphocytes, and centriacinar ducts, small ducts and islets of Langerhans in the pancreas. Further analysis in normal and diseased human pancreas show upregulated expression in diseased pancreas. Activation of caspase 2 in apoptosis was analyzed in two human cell lines in response to three different apoptotic stimuli: etoposide, staurosporine and Fas ligation. Immunoblotting with the monoclonal anti-caspase 2 antibody revealed no change in the quantity or molecular weight of the detected epitope during apoptosis induced with any of the three stimuli under conditions in which other caspases were clearly activated, caspase substrate cleavage occurred and the molecular changes of apoptosis were evident. It appears that caspase 2 is not critical for the induction of apoptosis in at least some cell types.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available