Noninvasive online biomass detector system for cultivation in shake flasks

Abstract

A novel online sensor system for noninvasive and continuous monitoring of cell growth in shake flasks is described. The measurement principle is based on turbidity measurement by detecting 180°‐scattered light and correlation to OD by nonlinear calibration models. The sensor system was integrated into a commercial shaking tablar to read out turbidity from below the shake flasks bottom. The system was evaluated with two model microorganisms, Escherichia coli K12 as prokaryotic and Saccharomyces cerevisiae as eukaryotic model. The sensor allowed an accurate monitoring of turbidity and correlation with OD600 ≤ 30. The determination of online OD showed relative errors of about 7.5% for E. coli K12 and 12% for S. cerevisiae. This matches the errors of the laborious offline OD and thus facilitates to overcome the drawbacks of the classical method as risk of contamination and decreasing volumes through sampling. One major challenge was to ensure a defined, nonvarying measurement zone as the rotating suspension in the shake flask forms a liquid sickle which circulates round the flasks inner bottom wall. The resulting alteration of liquid height above the sensor could be compensated by integration of an acceleration sensor into the tablar to synchronize the sensor triggering.

New perspectives in shake flask pH control using a 3D-printed control unit based on pH online measurement

“…In 2014 Scheper and Beutel et al proved its 9 biocompatibility and autoclavability for labware in use. Well plates and shake flask caps were 10printed and tested successfully with yeast cells and mammalian cells[24,25].…”

Monitoring of Microalgal Processes

“…Based on this idea, a novel online sensor system for noninvasive monitoring of biomass concentration by turbidity measurement in cultivation vessels has been developed [30]. Scattered light is measured through the vessel wall and correlated to optical density using nonlinear calibration models.…”

“…Other advantages offered by working at the laboratory scale are the optically clear cultivation medium prepared from laboratory-grade chemicals and clean water, and the transparent reactor walls enabling noninvasive optical measurement with sensors fixed on the outside. Such sensors can be employed not only for reading of optical density and fluorescence [17], of scattered light [30], or for advanced sensors as Raman spectroscopy [31] but also for physicochemical variables as pH, pO 2 , and pCO 2 with fluorescing sensor spots attached inside the reactor or shake flask [30] and read out by fiber optics placed on the outside. Microalgae also have specific advantages for sensor development in contrast to other microorganisms: microalgal cells are larger than bacteria and contain specific pigments absorbing in the visual range but not in the infrared [32], contain organelles small enough for Mie light scattering in the visible (VIS) and near-infrared (NIR) range [33], and the principal pigment, chlorophyll a, can be used as a probe of photosynthetic activity thanks to its fluorescence properties [34].…”

Application of an Online-Biomass Sensor in an Optical Multisensory Platform Prototype for Growth Monitoring of Biotechnical Relevant Microorganism and Cell Lines in Single-Use Shake Flasks

“…Several model microorganisms and cell lines representing aerobic and facultative anaerobic bacteria, fungi, yeasts and mammalian cells were chosen for the evaluation of the sensor. The technical design and the functional principle using an acceleration sensor have been described previously [ 1 ].…”

“…The focus lies on the evaluation of the biomass sensor based on backward light scattering. The application spectrum was expanded to four new organisms in addition to E. coli K12 and S. cerevisiae [ 1 ]. It could be shown that the sensor is appropriate for a wide range of standard microorganisms, e.g., L. zeae, K. pastoris, A. niger and CHO-K1.…”