The use of Raman spectroscopy for the live aseptic monitoring of stem cell differentiation

The characterisation of stem cell differentiation is currently performed using invasive/destructive methods, precluding cells from further use. There is an unmet need for techniques capable of characterising cells non-invasively in order to enhance the translation of stem cell-based therapies. Raman spectroscopy has previously been used to characterise cells and is non-invasive. However, almost all previous studies were not performed under aseptic conditions precluding any further use of those cells. The aim of this thesis was to determine whether Raman spectra could be acquired from the same stem cell cultures undergoing differentiation over time without compromising the sterility of these cell cultures and to identify aspects of the Raman spectra that may be used to predict eventual cell fate. Custom-built cell culture flasks with quartz windows (appropriate for acquiring Raman spectra) provided a platform for the maintenance of cell sterility whilst permitting the acquisition of Raman spectra comparable to those in the published literature. It was found that the osteogenic differentiation of dental pulp stem cells was associated with changes in protein content early in the process and prior to the end result being evident by the emergence of mineralised nodules. The adipogenic differentiation of adipose derived stem cells was found to be characterised by changes in lipid content throughout differentiation with the earliest detectable change after 3 days of differentiation. This is earlier than can typically be detected using currently established methods. Stem cell differentiation was also confirmed using conventional techniques including qRT-PCR and histology. Raman spectroscopy cannot provide the intricate detail of genetic/protein changes that occur as stem cells differentiate. However, this thesis demonstrates that stem cell fate can be monitored in an entirely non-invasive manner and could be applied to the scaled up production of differentiated stem cells for either clinical use or in a research setting.