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Small volume cell culture technology for the analysis of clonal heterogeneity in mammalian cell populations

University of British Columbia

The ability to culture individual cells provides a unique method to assess the heterogeneity of mammalian cell populations. However, there are many challenges when scaling down culture systems due to the complexity of re-creating a stimulating environment at the clonal level. Small volume culture systems such as integrated microfluidic platforms offer the potential to radically alter the throughput of clonal screening through the use of time-lapse imaging, dynamic stimulus control and economy of scale. In particular, the use of automated fluidic control allows for the characterization of single cells in a dynamic microenvironment similar to large-scale culture. This thesis describes how small volume cell culture practices such as the use of conditioned medium and microfluidic technology can be implemented to isolate large numbers of cells in small volumes and evaluate clonal populations under precise medium conditions. For a Chinese Hamster Ovary (CHO) cell system normal growth kinetics and specific productivity were sustained in small volumes. When exposed to conditioned medium from a parental CHO line, clones cultured at sub-mL scales matched the performance of large-scale cultures. A microfluidic bead assay was developed to detect Immunoglobulin G titers secreted from clones in nL volumes. The combination of microfluidic conditioned medium perfusion with the magnetic bead assay allowed for clonal productivity to be evaluated under simulated fed-batch conditions. Lastly, microfluidic cell culture was demonstrated on a human embryonic stem cell (hESC) system through the robust generation of colonies derived from single cells. hESCs propagated in the microfluidic system were observed to match the growth kinetics, marker expression and colony morphologies of larger cultures, while resolving response heterogeneity during differentiation induction. This thesis demonstrates how high-throughput, small volume culture systems can be used to screen clonal populations for therapeutic applications under complex culture conditions.

Text

2017-04-11

Attribution-NonCommercial-NoDerivatives 4.0 International

http://hdl.handle.net/2429/61186

Biomedical Engineering

University of British Columbia

UBCV

http://creativecommons.org/licenses/by-nc-nd/4.0/