Membrane‐aerated microbioreactor for high‐throughput bioprocessing

Abstract

A microbioreactor with a volume of microliters is fabricated out of poly(dimethylsiloxane) (PDMS) and glass. Aeration of microbial cultures is through a gas-permeable PDMS membrane. Sensors are integrated for on-line measurement of optical density (OD), dissolved oxygen (DO), and pH. All three parameter measurements are based on optical methods. Optical density is monitored via transmittance measurements through the well of the microbioreactor while dissolved oxygen and pH are measured using fluorescence lifetime-based sensors incorporated into the body of the microbioreactor. Bacterial fermentations carried out in the microbioreactor under well-defined conditions are compared to results obtained in a 500-mL bench-scale bioreactor. It is shown that the behavior of the bacteria in the microbioreactor is similar to that in the larger bioreactor. This similarity includes growth kinetics, dissolved oxygen profile within the vessel over time, pH profile over time, final number of cells, and cell morphology. Results from off-line analysis of the medium to examine organic acid production and substrate utilization are presented. By changing the gaseous environmental conditions, it is demonstrated that oxygen levels within the microbioreactor can be manipulated. Furthermore, it is demonstrated that the sensitivity and reproducibility of the microbioreactor system are such that statistically significant differences in the time evolution of the OD, DO, and pH can be used to distinguish between different physiological states. Finally, modeling of the transient oxygen transfer within the microbioreactor based on observed and predicted growth kinetics is used to quantitatively characterize oxygen depletion in the system.

Real‐time pH monitoring of industrially relevant enzymatic reactions in a microfluidic side‐entry reactor (μSER) shows potential for pH control

“…The importance of online monitoring has been described among the development needs for microbioreactors [40,41], and timecourse profiles of process parameters have been shown for fermentation [42] and stem cell culture [43]. Therefore, further applications of this approach are not only of interest for analytical and synthetic enzymatic reactions, enzyme stability studies or bioprocess development, but for a variety of biotechnological applications where continuous flow microreactors can be envisioned.…”

Microbioreactors for Bioprocess Development

“…Optical sensors were also used in the 6 mL miniature bioreactor reported by Lamping et a/. (2003), as well as in the microbioreactor described by Zanzotto et a/. (2004) Walther et al (1994Walther et al ( , 1999Walther et al ( , and 2000 developed a 3 mL continuous bioreactor with integrated biomass, pH, and temperature microelectronic sensors to investigate physiological and morphological properties of yeast cells in microgravity environments (in space lab).…”

“…A novel microchemostat system was recently reported by Balagadde et al (2005) Shaking is another way to improve gas-liquid oxygen transportation in bioreactors (Duetz et at., 2000;. Zanzotto et al (2004) provided oxygenation to cells growing in the microbioreactor by diffusion through a thin, gas-permeable PDMS membrane. In this reactor, PDMS membrane separates gas and liquid phases thereby eliminating the risk of contamination from air and cross-talk between adjacent microbioreactors.…”

“…Chapter Two addresses the fundamental issues in constructing a well-mixed, polymer-based, and instrumented batch microbioreactor as an extension of the membrane-aerated microbioreactor by Zanzotto et al (2004). Active mixing is achieved by using a mini magnetic stir bar driven by an external rotating magnetic field.…”

“…where Tm is the measured time constant of oxygen mass transfer, (Zanzotto et at., 2004) Global genomic expression assays using DNA microarrays permit the exploration of the cells transcriptional state under a wide range of physiological parameters. Microarray analysis has been widely applied in basic biological research as well as in specialized fields, such as drug screening, environmental testing, and clinical diagnosis (Debouck et al, 1999;Bodrossy et al, 2004).…”