Bioprocess Control in Microscale: Scalable Fermentations in Disposable and User-Friendly Microfluidic Systems

Abstract

BackgroundThe efficiency of biotechnological production processes depends on selecting the best performing microbial strain and the optimal cultivation conditions. Thus, many experiments have to be conducted, which conflicts with the demand to speed up drug development processes. Consequently, there is a great need for high-throughput devices that allow rapid and reliable bioprocess development. This need is addressed, for example, by the fiber-optic online-monitoring system BioLector which utilizes the wells of shaken microtiter plates (MTPs) as small-scale fermenters. To further improve the application of MTPs as microbioreactors, in this paper, the BioLector technology is combined with microfluidic bioprocess control in MTPs. To realize a user-friendly system for routine laboratory work, disposable microfluidic MTPs are utilized which are actuated by a user-friendly pneumatic hardware.ResultsThis novel microfermentation system was tested in pH-controlled batch as well as in fed-batch fermentations of Escherichia coli. The pH-value in the culture broth could be kept in a narrow dead band of 0.03 around the pH-setpoint, by pneumatically dosing ammonia solution and phosphoric acid to each culture well. Furthermore, fed-batch cultivations with linear and exponential feeding of 500 g/L glucose solution were conducted. Finally, the scale-up potential of the microscale fermentations was evaluated by comparing the obtained results to that of fully controlled fermentations in a 2 L laboratory-scale fermenter (working volume of 1 L). The scale-up was realized by keeping the volumetric mass transfer coefficient kLa constant at a value of 460 1/h. The same growth behavior of the E. coli cultures could be observed on both scales.ConclusionIn microfluidic MTPs, pH-controlled batch as well as fed-batch fermentations were successfully performed. The liquid dosing as well as the biomass growth kinetics of the process-controlled fermentations agreed well both in the microscale and laboratory scale. In conclusion, a user-friendly and disposable microfluidic system could be established which allows scaleable, fully controlled and fully monitored fermentations in working volumes below 1 milliliter.

Direct and indirect use of GFP whole cell biosensors for the assessment of bioprocess performances: Design of milliliter scale‐down bioreactors

“…Analysis of the extracellular proteome revealed that the most abundant proteins have a periplasmic origin (64% of this extracellular proteome is composed of periplasmic and outer membrane proteins). This observation is in accordance with the results of Funke et al (2011)38 who found, respectively, 65% of periplasmic proteins and 56% of outer membrane proteins with the W3110 Escherichia coli strain. However, some major cytoplasmic proteins have been identified, such as chaperonin GroE , chaperone DnaK , and elongation factor Tu.…”

“…Second, pH is not controlled in the mini‐bioreactors,33 potentially leading to the induction of some related regulation network linked with substrate starvation (i.e., general stress response pathway34–37). It must be noted that this technical issue could be corrected in the future by appropriate sensor‐actuator systems at the milliliter scale 23, 38. However, the media are buffered, so the maximum decrease in pH is 0.5 units (data not shown).…”

Quantification of the oxygen uptake rate in a dissolved oxygen controlled oscillating jet‐driven microbioreactor

“…In shaken systems, despite the significant advances in the monitoring of process variables, process control remains a limitation, making accurate determination of OUR in such devices challenging. In microtiter plates, this has been circumvented by applying feeding strategies to control pH and provide fed‐batch capabilities . Recently, several automated miniaturized systems based on plate formats have been commercialized, which have the ability to monitor and control in individual wells the pH, DO and temperature (e.g.…”

Anchoring high-throughput screening methods to scale-up bioproduction of siderophores

“…3B). Previously, other authors have shown correspondence between DO profiles and biomass production [29][30][31][32][33], suggesting that bacterial growth may be followed without the need for sampling or for interfering with the system. However, there were some discrepancies in the observed trends in biomass and DO in the case of M. smegmatis (Fig.…”