Be FAIR to your data

Wouldn’t it be great, if experimental data were findable wherever they were? If experimental data were accessible‚ regardless of the storage place and format? If experimental data were interoperable independent of the author or its origin? If experimental data were reusable for further analysis without experimental repetition? The current state of the art of data acquisition in the laboratory is very diverse. A lot of different devices are used, analogue as well as digital ones. Usually all experimental setups and observations are summarized in a handwritten lab notebook, independently from digital or analogue sources. To change the actual and common way of laboratory data acquisition into a digital and modern one, electronic lab notebooks can be used. A challenge of science is to facilitate knowledge discovery by assisting humans and machines in their discovery of scientific data and their associated algorithms and workflows. FAIR describes a set of guiding principles to make data Findable, Accessible, Interoperable, and Reusable

Analyse und Optimierung eines industriellen Biotransformationsprozesses zur Herstellung von Tryptophan

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Establishment of a Perfusion Process with Antibody-Producing CHO Cells Using a 3D-Printed Microfluidic Spiral Separator with Web-Based Flow Control

Monoclonal antibodies are increasingly dominating the market for human therapeutic and diagnostic agents. For this reason, continuous methods—such as perfusion processes—are being explored and optimized in an ongoing effort to increase product yields. Unfortunately, many established cell retention devices—such as tangential flow filtration—rely on membranes that are prone to clogging, fouling, and undesirable product retention at high cell densities. To circumvent these problems, in this work, we have developed a 3D-printed microfluidic spiral separator for cell retention, which can readily be adapted and replaced according to process conditions (i.e., a plug-and-play system) due to the fast and flexible 3D printing technique. In addition, this system was also expanded to include automatic flushing, web-based control, and notification via a cellphone application. This set-up constitutes a proof of concept that was successful at inducing a stable process operation at a viable cell concentration of 10–17 × 106 cells/mL in a hybrid mode (with alternating cell retention and cell bleed phases) while significantly reducing both shear stress and channel blockage. In addition to increasing efficiency to nearly 100%, this microfluidic device also improved production conditions by successfully separating dead cells and cell debris and increasing cell viability within the bioreactor

Employing QbD strategies to assess the impact of cell viability and density on the primary recovery of monoclonal antibodies

Quality by Design (QbD) is one of the most important tools for the implementation of Process Analytical Technology (PAT) in biopharmaceutical production. For optimal characterization of a monoclonal antibody (mAb) upstream process a stepwise approach was implemented. The upstream was divided into three process stages, namely inoculum expansion, production, and primary recovery, which were investigated individually. This approach enables analysis of process parameters and associated intermediate quality attributes as well as systematic knowledge transfer to subsequent process steps. Following previous research, this study focuses on the primary recovery of the mAb and thereby marks the final step toward a holistic characterization of the upstream process. Based on gained knowledge during the production process evaluation, the cell viability and density were determined as critical parameters for the primary recovery. Directed cell viability adjustment was achieved using cytotoxic camptothecin in a novel protocol. Additionally, the cell separation method was added to the Design of Experiments (DoE) as a qualitative factor and varied between filtration and centrifugation. To assess the quality attributes after cell separation, the bioactivity of the mAb was analyzed using a cell-based assay and the purity of the supernatant was evaluated by measurement of process related impurities (host cell protein proportion, residual DNA). Multivariate data analysis of the compiled data confirmed the hypothesis that the upstream process has no significant influence on the bioactivity of the mAb. Therefore, process control must be tuned towards high mAb titers and purity after the primary recovery, enabling optimal downstream processing of the product. To minimize amounts of host cell proteins and residual DNA the cell viability should be maintained above 85% and the cell density should be controlled around 15 × 106 cells/ml during the cell removal. Thereby, this study shows the importance of QbD for the characterization of the primary recovery of mAbs and highlights the useful implementation of the stepwise approach over subsequent process stages

Implementation of QbD strategies in the inoculum expansion of a mAb production process

The quality by design approach was introduced to the biopharmaceutical industry over 15 years ago. This principle is widely implemented in the characterization of monoclonal antibody production processes. Anyway, the early process phase, namely the inoculum expansion, was not yet investigated and characterized for most processes. In order to increase the understanding of early process parameter interactions and their influence on the later production process, a risk assessment followed by a design of experiments approach was conducted. The DoE included the critical parameters methotrexate (MTX) concentration, initial passage viable cell density and passage duration. Multivariate data analysis led to mathematical regression models and the establishment of a designated design space for the studied parameters. It was found that the passage duration as well as the initial viable cell density for each passage during the inoculum expansion have severe effects on the growth rate and viability of the early process phase. Furthermore, the variations during the inoculum expansion directly influenced the production process responses. This carry-over of factor effects highlights the crucial impact of early process failures and the importance of process analysis and control during the first part of mAb production processes. © 2020 The Authors. Engineering in Life Sciences published by Wiley-VCH Gmb

NIR-spectroscopy for bioprocess monitoring & control

The Ingold port adaption of a free beam NIR spectrometer is tailored for optimal bioprocess monitoring and control. The device shows an excellent signal to noise ratio dedicated to a large free aperture and therefore a large sample volume. This can be seen particularly in the batch trajectories which show a high reproducibility. The robust and compact design withstands rough process environments as well as SIP/CIP cycles. Robust free beam NIR process analyzers are indispensable tools within the PAT/QbD framework for realtime process monitoring and control. They enable multiparametric, non-invasive measurements of analyte concentrations and process trajectories. Free beam NIR spectrometers are an ideal tool to define golden batches and process borders in the sense of QbD. Moreover, sophisticated data analysis both quantitative and MSPC yields directly to a far better process understanding. Information can be provided online in easy to interpret graphs which allow the operator to make fast and knowledge-based decisions. This finally leads to higher stability in process operation, better performance and less failed batches

Development and characterization of a fiber optical fluorescence sensor for the online monitoring of biofilms and their microenvironment

The growth of microorganisms on surfaces and interfaces as a biofilm is very common and plays important role in various areas such as material science, biomedicine, or waste treatment among others. Due to their inhomogeneous structure and the variance in the microorganism consortium, the analysis of biofilms represents a significant challenge. An online fluorescence sensor was developed that is able to measure the most important biological fluorophores (proteins, nicotinamide adenine dinucleotide, and flavin) in a noninvasive manner in biofilms, e.g. in bioelectrochemical applications. The sensor gives the opportunity to continuously draw conclusions on the metabolic state of the biofilm. The developed sensor has a diameter of 1 mm at the sensor tip and can be moved on and into the biofilm surface. In the first experiment, the measuring range of the sensor and the long‐term stability could be determined and the system applicability was confirmed. In addition, measurements in biofilm‐like structures could be performed. The formation of a wastewater‐based biofilm was monitored using the developed sensor, demonstrating the functionality of the sensor in a proof‐of‐principle experiment

Stress‐induced increase of monoclonal antibody production in CHO cells

Monoclonal antibodies (mAbs) are of great interest to the biopharmaceutical industry due to their widely used application as human therapeutic and diagnostic agents. As such, mAb require to exhibit human‐like glycolization patterns. Therefore, recombinant Chinese hamster ovary (CHO) cells are the favored production organisms; many relevant biopharmaceuticals are already produced by this cell type. To optimize the mAb yield in CHO DG44 cells a corelation between stress‐induced cell size expansion and increased specific productivity was investigated. CO(2) and macronutrient supply of the cells during a 12‐day fed‐batch cultivation process were tested as stress factors. Shake flasks (500 mL) and a small‐scale bioreactor system (15 mL) were used for the cultivation experiments and compared in terms of their effect on cell diameter, integral viable cell concentration (IVCC), and cell‐specific productivity. The achieved stress‐induced increase in cell‐specific productivity of up to 94.94.9%–134.4% correlates to a cell diameter shift of up to 7.34 μm. The highest final product titer of 4 g/L was reached by glucose oversupply during the batch phase of the process

Digitale Zwillinge in der Bioprozesstechnik – Chancen und Möglichkeiten

The digitalization of processes is one of the currently dominating topics and missions in both industrial production and scientific research. In biotechnology, these efforts have enormous potential to optimize existing bioprocesses or to develop new bioprocesses for upcoming challenges. In this review, the state-of-art of digitalization in bioprocess engineering is considered, the terms digital twin and digital shadow are characterized, and an outlook on future Lab 4.0 concepts is given.Die Digitalisierung von Prozessen ist eines der aktuell dominierenden Themen und Missionen sowohl in der industriellen Produktion als auch der wissenschaftlichen Forschung. In der Biotechnologie haben diese Bemühungen ein enormes Potenzial, um existierende Bioprozesse zu optimieren oder neue Bioprozesse für zukünftige Herausforderungen zu entwickeln. In diesem Übersichtsbeitrag wird der aktuelle Stand der Digitalisierung in der Bioprozesstechnik beleuchtet, die Begriffe digitaler Zwilling und digitaler Schatten charakterisiert sowie ein Ausblick auf zukünftige Lab 4.0-Konzepte gegeben