UBC Theses and Dissertations
Assessment of the onset and progress of apoptosis via Raman spectroscopy Rangan, Shreyas
Apoptotic and necrotic cell death is ultimately the cause of productivity loss in bioreactors used to produce therapeutic proteins. This study investigates the suitability of Raman spectroscopy to detect the onset and types of cell death in Chinese Hamster Ovary (CHO) cells - the most widely used cell type for therapeutic protein production. Apoptotic, necrotic or autophagic CHO cells producing tissue plasminogen activator were compared to uninduced cultures using Raman spectroscopy and principal component analysis (PCA). A fingerprint region was identified where several peaks change in the course of cell death. Further, uninduced cells were compared to cells sorted at different stages of apoptosis, in order to establish how early the onset of apoptosis could be detected. These results move past what has been described in literature, as we have shown that apoptosis-induced cells that score as viable in conventional apoptosis assays appear to have an altered biochemical composition compared to uninduced cells. Cells from different stages of fed-batch cultures were compared, and the results showed that Raman spectroscopy can be used to monitor the progress of a fed-batch culture. However, further work is required to elucidate the onset of apoptosis in fed-batch cultures. Future goals include assessing different inducers of apoptosis in order to construct a "library" of biochemical changes during apoptotic cell death, and developing automated classification models such as support vector machines to classify cell death types.
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