Biology subjects

Neubauer, P.

Publications and source records attributed to Neubauer, P..

6 recordsLinked to original sources

bioRxiv2023

Automation of Experimental Workflows for High Throughput Robotic Cultivations

Process systems engineering methods and tools have been difficult to apply in bioprocess engineering, mainly due to the high complexity of biological systems and the low reproducibility of the experiments. High throughput robotic cultivation platforms in combination with computational tools for experimental design, resource scheduling, and operation, are rapidly gaining popularity. One important contribution being the generation of data in high throughput needed to overcome this lack of data with high information content and the worrying reproducibility crisis in life sciences. In this work, directed acyclic graphs are used to represent, manage and track all experimental workflows in a robotic platform. They support data provenance and enable traceability and reproducibility of workflows in robotic facilities. The experimental workflows are automated using Apache Airflow enabling to manage all necessary steps for fed-batch cultivations, including sampling, sample transport by a mobile robot, feed additions, data collection, storage in a SQL database and model fitting. The added value of this system is demonstrated in scale-down experiments, where E. coli BL21 (DE3), producing elastin like proteins, exhibits robustness towards glucose oscillations that mimic industrial cultivation conditions.

bioengineering↗

bioRxiv2023

Modeling of enzyme-mediated glucose release to facilitate continuous feed in miniaturized cultivations

When striving for maximal throughput at minimal volumes while cultivating close to industrial conditions, simple and robust feeding strategies offer important advantages. Enzyme-mediated glucose cleavage from dextrin is an easy way of imitating continuous fed-batch in the small scale, with no complex equipment required. While the release rate - and thus the feed rate - can be controlled by adapting the enzyme concentration, it strongly depends on the concentration of the involved substances and the environmental conditions. Thus, it is a challenge to use the technology for controlling the specific growth rate, as it is commonly done with feed pumps. For solving this problem, we present here a mathematical model that extends simple Michaelis-Menten kinetics by considering different substrate fractions and can be applied to control the glucose release rate even in high throughput experiments. The fitted model was used during automated microbial cultivations to control the growth rate in quasi-continuous fed-batch processes and to realize different exponential growth rates by intermittent additions of enzyme and dextrin by a liquid handling robot system. We thus present an approach for defined biocatalytically controlled glucose supply of small-scale systems, where - if at all - continuous feeding was only possible with low accuracy or high technical efforts until now.

bioengineering↗

bioRxiv2022

Automated bioprocess feedback operation in a high throughput facility via the integration of a mobile robotic lab assistant

Biotechnological processes development is challenging due to the sheer variety of process parameters. For efficient upstream development parallel cultivation systems have proven to reduce costs and associated timelines successfully, while offering excellent process control. However, the degree of automation of such small scale systems is comparably low and necessary sample analysis requires manual steps. Although the subsequent analysis can be performed in a high-throughput manner, the integration of analytic devices remains challenging. Especially, when cultivation and analysis laboratories are spatially separated. Mobile robots offer a potential solution, but the implementation in research laboratories is not widely adopted. Our approach demonstrates the integration of a small scale cultivation system into a liquid handling station for an automated sample procedure. The samples are transferred via a mobile robotic lab assistant and subsequently analysed by a high-throughput analyzer. The process data is stored in a centralized database. The mobile robotic workflow guarantees a flexible solution for device integration and facilitates automation. Restrictions regarding spatial separation of devices are circumvented, enabling a modular platform throughout different laboratories. The presented cultivation platform is evaluated based on industrial relevant E. coli BW25113 high cell density fed-batch cultivation. Here its suitability for accelerating bioprocess development is proven. The necessary magnesium addition for reaching high cell densities in mineral salt medium is automated via a feedback operation loop. The feedback operation loop demonstrates the possibility for advanced control options. This study sets the foundation for a fully integrated facility with different cultivation scales sharing the same data infrastructure, where the mobile robotic lab assistant physically connects the devices.

bioengineering↗

bioRxiv2021

Optimal operation of parallel mini-bioreactors in bioprocess development using multi-stage MPC

Bioprocess development is commonly characterized by long development times, especially in the early screening phase. After promising candidates have been pre-selected in screening campaigns, an optimal operating strategy has to be found and verified under conditions similar to production. Cultivating cells with pulse-based feeding and thus exposing them to oscillating feast and famine phases has shown to be a powerful approach to study microorganisms closer to industrial bioreactor conditions. In view of the large number of strains and the process conditions to be tested, high-throughput cultivation systems provide an essential tool to sample the large design space in short time. We have recently presented a comprehensive platform, consisting of two liquid handling stations coupled with a model-based experimental design and operation framework to increase the efficiency in High Throughput bioprocess development. Using calibrated macro-kinetic growth models, the platform has been successfully used for the development of scale-down fed-batch cultivations in parallel mini-bioreactor systems. However, it has also been shown that parametric uncertainties in the models can significantly affect the prediction accuracy and thus the reliability of optimized cultivation strategies. To tackle this issue, we implemented a multi-stage Model Predictive Control (MPC) strategy to fulfill the experimental objectives under tight constraints despite the uncertainty in the parameters and the measurements. Dealing with uncertainties in the parameters is of major importance, since constraint violation would easily occur otherwise, which in turn could have adverse effects on the quality of the heterologous protein produced. Multi-stage approaches build up scenario tree, based on the uncertainty that can be encountered and computing optimal inputs that satisfy the constrains despite of such uncertainties. Using the feedback information gained through the evolution along the tree, the control approach is significantly more robust than standard MPC approaches without being overly conservative. We show in this study that the application of multi-stage MPC can increase the number of successful experiments, by applying this methodology to a mini-bioreactor cultivation operated in parallel.

bioengineering↗

bioRxiv2020

Suspension of A Point-Mass-Loaded Filament in Non-uniform Flows: Passive Dynamics of a Ballooning Spider

Spiders utilize their fine silk fibres for their aerial dispersal, known as ballooning. With this method, spiders can disperse hundreds of kilometres, reaching as high as 4.5 km. However, the passive dynamics of a ballooning model (a highly flexible filament and a spider body at the end of it) are not well understood. The previous study (Rouse model: without taking into account anisotropic drag of a fibre) suggested that the flexible and extendible fibres reduce the settling speed of the ballooning model in homogeneous turbulence. However, the exact cause of the reduction of the settling speed is not explained and the assumed isotropic drag of a fibre is not realistic in the low Reynolds number flow. Here we introduce a bead-spring model that takes into account the anisotropic drag of a fibre to investigate the passive behaviour of the ballooning model in the various non-uniform flows (a shear flow, a periodic vortex flow field and a homogeneous turbulent flow). For the analysis of the wide range of parameters, we defined a dimensionless parameter, which is called a ballooning number. The ballooning number means the ratio of Stokes fluid-dynamic force on a fibre by the non-uniform flow field to the gravitational force of a body at the end of the fibre. Our simulation shows that the settling speed of the present model in the homogeneous turbulent flows shows the biased characters of slow settling as the influence of the turbulent flow increases. The causes of this slow settling are investigated by simulating it in a wide range of shear flows. We revealed that the cause of this is the drag anisotropy of the filament structure (spider silk). In more detail, the cause of reduced settling speed lies not only in the deformed geometrical shape of the ballooning silk but also in its generation of fluid-dynamic force in a non-uniform flow (shear flow). Additionally, we found that the ballooning structure could become trapped in a vortex flow. This seemed to be the second reason why the ballooning structure settles slowly in the homogeneous turbulent flow. These results can help deepen our understanding of the passive dynamics of spiders ballooning in the atmospheric boundary layer.

biophysics↗

bioRxiv2017

An Observational Study of Ballooning in Large Spiders: Nanoscale Multi-Fibres Enable Large Spiders’ Soaring Flight

The physical mechanism of aerial dispersal of spiders, \"ballooning behavior,\" is still unclear because of the lack of serious scientific observations and experiments. Therefore, as a first step in clarifying the phenomenon, we studied the ballooning behavior of relatively large spiders (heavier than 5 mg) in nature. Additional wind tunnel tests to identify ballooning silks were implemented in the laboratory. From our observation, it seems obvious that spiders actively evaluate the condition of the wind with their front leg (leg I) and wait for the preferable wind condition for their ballooning takeoff. In the wind tunnel tests, as yet unknown physical properties of ballooning fibers (length, thickness and number of fibers) were identified. Large spiders, 16-20 mg Xysticus species, spun 50 to 60 nanoscale fibers, with a diameter of 121 to 323 nm. The length of these threads was 3.22 {+/-} 1.31 m (N = 22). These physical properties of ballooning fibers can explain the ballooning of large spiders with relatively light updrafts, 0.1-0.5 m s-1, which exist in a light breeze of 1.5-3.3 m s-1. Additionally, in line with previous research on turbulence in atmospheric boundary layers and from our wind measurements, it is hypothesized that spiders use the ascending air current for their aerial dispersal, the \"ejection\" regime, which is induced by hairpin vortices in the atmospheric boundary layer turbulence. This regime is highly correlated with lower wind speeds. This coincides well with the fact that spiders usually balloon when the wind speed is lower than 3 m s-1.

biophysics↗