Development of a novel bioreactor and systems for suspension cell culture in biopharmaceutical production
This is the data linked to the Doctoral thesis titled 'Development of a novel bioreactor and systems for suspension cell culture in biopharmaceutical production' (2021).
Mammalian cells offer superior cellular machinery for the production of complex biological products. These cells provide proper post-translational processing machinery for recombinant protein expression to acquire the desired folding for optimal activity. With this advantage, mammalian cells have become the preferred choice for the production of biologicals, e.g. biotherapeutic proteins, monoclonal antibodies, and vaccines. These cells may grow either attached to a solid surface for their growth (adherent cells) or, where adapted, as suspension cultures. In order to grow these cells efficiently, a bioreactor is therefore required.
In this study, CHO cells have been successfully adapted to suspension in serum-free conditions using the slow weaning of serum method and propagated in the HTB whereas Vero cells have been adapted successfully to serum-free media in adherent conditions. Attempt to suspend Vero cells based on literature using the weaning method remains timeous. Therefore, an alternative approach was explored using an anti-cancer drug (PAN) which is known to suppress the expression of integrin (cell adhesion receptors). The expectations from this approach was that the suppression of integrin would allow cells to detach and grow as a suspended culture (Krishnamurti et al., 2001). The results indicated that the anti-cancerous drug may have modulated the structure and function of the integrin which resulted in dislodging of the cells from the surface and form clumps which were viable for a week in suspension culture without increase in cell density. The viability of the cell clumps and few suspended cells were tested by re-seeding of these cells back to tissue culture (TC) flasks in serum-containing media without the presence of PAN. The culture in the TC flask regained confluency in the 2-3 day which confirms the viability of the cells and the likeliness of integrin re-modulating itself in the absence of PAN. As the suspended Vero cells did not grow, they were not tested for growth in the HTB.
To investigate the biological activity of these Vero cells, Isothermal microcalorimetry was used to evaluate the heat generation profile of the Vero cells quantitatively before and after drug treatment. The heat flow data (metabolic heat) from the treated and normal cells showed a distinct decrease in the heat generation profile which indicated that the treated cells were viable but not as active as the normal (non-treated) cells. It was evident from the heat flow data obtained for the PAN-treated Vero cells (-0.13 µW) from that of non-treated cells (13.12 µW) and thereafter when PAN-treated Vero cells regrown in serum-containing media, they regain their metabolic activities which were indicated by their heat flow values as positive control (9.30 µW), 100 µg mL-1 (10.12 µW), 200 µg mL-1 (10.18 µW), and 250 µg mL1 (9.15 µW).
2. Literature Review
3. Approach and Methodology
4. Designing, fabrication and abiotic characterisation of HTB
5. Biotic characterization of HTB
6. Exploiting puromycin aminonucleoside (PAN) to bring strongly adherent Vero cells into suspension
7. Investigating isothermal microcalorimetric profile of post-drug-treated Vero cells for the suspension culture