Finding the CHOsen one – Optimising biopharmaceutical-cell line selection via advanced molecular microscopy techniques

Thomas, Rhodri 2024. Finding the CHOsen one – Optimising biopharmaceutical-cell line selection via advanced molecular microscopy techniques. PhD Thesis, Cardiff University. Item availability restricted.

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Abstract

The Chinese Hamster Ovary (CHO) cell line is the dominant expression host used in industrial scale production of monoclonal antibodies (mAbs). A significant problem when developing cell lines is clonal instability, whereby production output/quality drops over the timescale needed to scale up to bioreactor quantity. What is needed are better predictors of cell line stability in order to improve eventual cell line selection. The underlying hypothesis explored within this thesis is whether stability phenotypes of cell lines were reflected in morphological characteristics that would aid in selection of candidate cell lines for long term culture. This was assessed through three approaches. First, a direct cell labelling technique using live-cell fluorescent dyes explored organelles and cell structures associated with the secretory pathway. Second, a fluorescence based live-cell assay examined cargo flow through the secretory pathway. Finally, non-invasive, chemically specific CARS microscopy was explored to understand whether chemical spectra could be identified that would predict stability. Each of the methodologies were developed on a core panel of adherent cell types, then adapted to the CHO cell lines. Fluorescence profiling identified and characterised a panel of markers, that ultimately lacked the ability to separate CHO phenotypes. The secretory transport assay was confirmed as an assay capable of quantitatively identifying both defects within cell lines, and differences between cell lines, however the challenges of image capture/analysis revealed that throughput, sample movement and the long timecourse of experimentation would hinder translation into an industrial setting despite its initial promise. These problems were also highlighted within the CARS approach, where cell specific chemical spectra could be identified, but not associated with cell phenotypes. For each case it is discussed where challenges arose in the methodology, and where this valuable intersection of cellular biology, optical imaging, and data science can head next.

Item Type:	Thesis (PhD)
Date Type:	Completion
Status:	Unpublished
Schools:	Schools > Biosciences
Subjects:	Q Science > Q Science (General)
Date of First Compliant Deposit:	24 March 2025
Date of Acceptance:	24 March 2025
Last Modified:	24 Mar 2025 13:46
URI:	https://orca.cardiff.ac.uk/id/eprint/177105

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