Immobilization of transgenic plant cells towards bioprinting for production of a recombinant biodefense agent

Transgenic rice cells (Oryza sativa) producing recombinant butyrylcholinesterase (BChE) as a prophylactic/therapeutic against organophosphate nerve agent poisoning, cocaine toxicity, and neurodegenerative diseases like Alzheimer's were immobilized in a polyethylene glycol-based hydrogel. The cells were sustained for 14 days in the semi-solid matrix, undergoing a growth phase from days 0-6, a BChE production phase in sugar-free medium from days 6-12, and a growth/recovery phase from days 12-14. Throughout this period, the cells maintained similar viability to those in suspension cultures and displayed analogous sugar consumption trends. The rice cells in the hydrogel also produced a significant amount of active BChE, comparable to the levels produced in liquid cultures. A considerable fraction of this BChE was secreted into the media, allowing for easier product separation. To the best of our knowledge, this proof-of-concept is the first report of immobilization of recombinant plant cells for continuous production of high-value heterologous proteins. This work serves as a foundation for further investigation towards plant cell bioprinting and the development of a simple, efficient, robust, modular, and potentially field-deployable bioreactor system for the manufacture of biologics.