The use of Process Analytical Technologies to examine the viability of CHO cells

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Title: The use of Process Analytical Technologies to examine the viability of CHO cells
Authors: Bergin, Adam
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Date: 2022
Online since: 2022-11-07T16:01:06Z
Abstract: The viability of mammalian cells is primarily tested by dye exclusion assays to examine the integrity of the outer membrane. Precursor events to the onset of cell death are detectable using a combination of online and offline technologies. This work explores the use of dielectric spectroscopy and impedance flow cytometry to characterize changes in the biophysical properties of cells as they progress through batch cultures. At-line single cell imaging was examined in tandem with these methods to prove further insight into the identification of morphological changes in the cell culture. This information was collated to better understand at what point cells can no longer be classified as recoverable prior to the loss in membrane integrity. Autophagic activity such as the increased presence of lysosomes was identified using digital holographic imaging. An earlier decline in the online capacitance signal relative to offline counts occurred in tandem with the onset of autophagy due the shifting dynamics of the cell population. Schwan modelling gave insight on the changes in the bulk membrane capacitance and intracellular conductivity of the cells during this period. Single cell impedance measurements were used to examine the population dynamics with greater accuracy. Opacity and phase parameters were derived at suitable frequencies and compared to the online models. Multifrequency data from the capacitance probe proved useful in the identification of apoptotic activity which followed autophagy. The Cole-Cole a and critical frequency of the changing ß-dispersion curve properties were examined relative to these starvation events. A feeding strategy was employed to delay the onset of autophagy in batch cultures, through the introduction of amino acids. Controlled refeeding experiments were shown to affect both the presence of lysosomes and shifts in opacity trends, suggesting that cells could be recovered during autophagy. The effects of such a feed on the online modelling data was examined to see if a real time parameter from the multifrequency trends could be used as an indicator for culture refeeding.
Type of material: Doctoral Thesis
Publisher: University College Dublin. School of Chemical and Bioprocess Engineering
Qualification Name: Ph.D.
Copyright (published version): 2022 the Author
Keywords: CapacitanceImpedanceImagingAutophagy
Language: en
Status of Item: Peer reviewed
This item is made available under a Creative Commons License:
Appears in Collections:Chemical and Bioprocess Engineering Theses

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