Biochemical quantification to assess raman-based inferences of t-cell composition of DNA, RNA, and proteins

The development of T-cell therapies has resulted in major advancements in cancer immunotherapies. Raman spectroscopy has the potential to provide a noninvasive and optical means to monitor therapeutic T-cell manufacturing processes by assessing changes in cellular macromolecular composition, even at the single-cell level. This could serve as an additional means of validating the consistency of T-cell manufacturing for therapy, particularly as more artificial stem-cell-derived T-cells are produced. However, overlaps in Raman spectral peaks introduce ambiguity in identifying distinct macromolecular components in T-cells, and thus, complicates their quantification. To validate the inferences from Raman measurements, I aimed to accurately quantify the T-cell content of DNA, RNA, and proteins using conventional biochemical assays. In order to do this, I maximized nucleic acid yield from cells by reducing cell input to a silica-based binding method and measured total protein content through a colorimetric technique and another promising approach that utilized the nitrogen mass measured from cell combustion. These methods should now enable the refinement of Raman spectroscopy quantification methods and the selection of more reliable spectral biomarkers of T-cell composition for process analytical purposes.