UBC Theses and Dissertations
Cell death dynamics monitoring using Raman micro-spectroscopy Karimbabanezhadmamaghani, Pooya
Biopharmaceuticals play a crucial role in curing diseases like Cancer and diabetes. Bioreactors are the heart of the industry. Cell losses due to cell death such as apoptosis and necrosis in the bioreactor decreases production efficiency and subsequently increases the cost of production. Furthermore, the study of apoptosis and necrosis cell death mechanisms has a great scientific and clinical importance in cancer therapy. In this project, Raman micro-spectroscopy is used to study apoptosis and necrosis in Chinese’s Hamster Ovary (CHO) cells that are one the main host cell lines used in the production of biopharmaceuticals. Apoptosis and Necrosis were induced in CHO cells using camptothecin and oxygen and glucose deprivation. The changes in the chemical composition of these enriched apoptotic and necrotic cell cultures were then analyzed using Raman spectroscopy which revealed novel biological concepts of the cell death process. Moreover, highly distinguishing Raman characteristics were identified for each death mode. These observations made by Raman spectroscopy were confirmed using a broad range of conventional and advanced biological assays in the field ranging from FACS analysis and fluorescent dyes to fluorescence microscopy. Studying Raman Spectra gave a clear image about DNA, RNA and Protein level changes during the process of apoptosis and necrosis in CHO cells. Using Principle Components Analysis (PCR) enabled viable, necrotic, early and late apoptotic populations to be clearly distinguished. This technology may provide the basis for the development of a non-invasive probe to monitor and predict cell death in bioreactor cultures in real-time and possibly allow cultures to avoid entering the cell death phase. In addition, the vast majority of cancer treatment methods involve cell death and apoptosis and, therefore, improving our knowledge about the biology of cell death will help support and advance research and treatment in this area.
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Attribution-NonCommercial-NoDerivs 2.5 Canada