An integrated microfluidic device for continuous bioprocessing

Abstract

Microfluidic devices have been increasingly used as tools to accelerate the development of new biomanufacturing methods, given their ability to test a large set of variables in a short time, with minimal reagent consumption. However, while being able to expedite a bioprocess design, the modular aspect of these microfluidic devices has not been systematically explored, with most applications focusing on single unitary operations. Integrated continuous bioprocessing has the potential to reduce cost, optimize yield and reduce footprint as compared to batch operation. In this work an integrated microfluidic device for continuous processing is developed and qualitatively shows connected different unit operations able to produce GFP in microbioreactor, extract out of cells in lysis module and finally concentrate in ATPs module.The integrated system developed herein consists of a micro-chemostat module to produce a target protein in continuous mode, a chemical cell lysis module for protein release and a module to purify and concentrate the protein using aqueous two-phase systems (ATPS). In this way, it is possible to screen multiple conditions for each operation and evaluate their combined effect on the final product. A recombinant Escherichia coli (E. coli) strain producing green fluorescent protein (GFP) was used as a model system, which allows for GFP production, lysis efficiency and partition coefficient to be evaluated by fluorescence microscopy. Experiments were done for over a week by continuously supplying medium and monitoring the GFP production by fluorescence intensity. A chemical lysis solution B-PER® was used for continuous lysis and afterwards the extracted GFP was clarified and concentrated in the PEG-rich phase of a PEG/phosphate ATPS with a partition coefficient of 2. As the integrated device is working in continuous mode it could be integrated with further downstream processing modules.