Cross-section perimeter is a suitable parameter to describe the effects of different baffle geometries in shaken microtiter plates

Abstract

BackgroundBiotechnological screening processes are performed since more than 8 decades in small scale shaken bioreactors like shake flasks or microtiter plates. One of the major issues of such reactors is the sufficient oxygen supply of suspended microorganisms. Oxygen transfer into the bulk liquid can in general be increased by introducing suitable baffles at the reactor wall. However, a comprehensive and systematic characterization of baffled shaken bioreactors has never been carried out so far. Baffles often differ in number, size and shape. The exact geometry of baffles in glass lab ware like shake flasks is very difficult to reproduce from piece to piece due to the hard to control flow behavior of molten glass during manufacturing. Thus, reproducibility of the maximum oxygen transfer capacity in such baffled shake flasks is hardly given.ResultsAs a first step to systematically elucidate the general effect of different baffle geometries on shaken bioreactor performance, the maximum oxygen transfer capacity (OTRmax) in baffled 48-well microtiter plates as shaken model reactor was characterized. This type of bioreactor made of plastic material was chosen, as the exact geometry of the baffles can be fabricated by highly reproducible laser cutting. As a result, thirty different geometries were investigated regarding their maximum oxygen transfer capacity (OTRmax) and liquid distribution during shaking. The relative perimeter of the cross-section area as new fundamental geometric key parameter is introduced. An empirical correlation for the OTRmax as function of the relative perimeter, shaking frequency and filling volume is derived. For the first time, this correlation allows a systematic description of the maximum oxygen transfer capacity in baffled microtiter plates.ConclusionsCalculated and experimentally determined OTRmax values agree within ± 30% accuracy. Furthermore, undesired out-of-phase operating conditions can be identified by using the relative perimeter as key parameter. Finally, an optimum well geometry characterized by an increased perimeter of 10% compared to the unbaffled round geometry is identified. This study may also assist to comprehensively describe and optimize the baffles of shake flasks in future.

Quasi-continuous parallel online scattered light, fluorescence and dissolved oxygen tension measurement combined with monitoring of the oxygen transfer rate in each well of a shaken microtiter plate

“…3a–d, m–p). The general trend of decreasing OTR max with increasing filling volume is in agreement with other studies [23, 24]. The oxygen limitation occurs earlier with increasing filling volumes (700 µL: 9.9 h; 1400 µL: 9.1 h).…”

“…4 with an empirical correlation which was presented by Lattermann et al [23]. The empirical correlation [23] is illustrated as black line and the dashed black lines represent an error range of ±15%. For each filling volume, the maximum measured OTR values (≙OTR max ) for parallel cultures were averaged.…”

“…Already in 1965, it was shown that oxygen-limited conditions during cultivation can affect microbial growth and product formation [22]. Lattermann et al [23] presented an empirical correlation to estimate and compare the maximum oxygen transfer capacity (OTR max ) in MTPs with different well shapes. Besides a relative perimeter, which considers different well geometries, the shaking frequency (n) and the filling volume (V L ) are used in the empirical correlation for OTR max assessments.…”

Respiration activity monitoring system for any individual well of a 48-well microtiter plate

“…To increase the experimental throughput, the replicate number is later reduced to eight and three replicates and lastly singlet cultivations were performed. According to Lattermann et al [30], the predicted maximum oxygen transfer capacities are 10 mmol/l/h for the plant cell suspension culture and 45 mmol/l/h for the cultivation of H. polymorpha and E. coli at the applied cultivation conditions. These capacities are smaller compared to a typical stirred tank reactor system.…”

“…These capacities are smaller compared to a typical stirred tank reactor system. But considering different well geometries and shaking parameters, the resulting increased maximum oxygen transfer capacities can be comparable to stirred tank systems [30–32]. Based on the literature [16, 28, 33, 34], all cultivations of this section are expected to be oxygen unlimited.…”