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Mammalian cell culture systems produce high-value biologics, such as monoclonal antibodies, which are increasingly being used clinically. A complete framework that interlinks model-based design of experiments (DOE) and model-based control and optimization to the actual industrial bioprocess could assist experimentation, hence reducing costs. However, high fidelity models have the inherent characteristic of containing a large number of parameters, which is further complicated by limitations in the current analytical techniques, thus resulting in the experimental validation of merely a small number of parameters. Sensitivity analysis techniques can provide valuable insight into model characteristics. Traditionally, the application of sensitivity analysis on models of biological systems has been treated more or less as a black box operation. In the present work, we elucidate the aspects of sensitivity analysis and identify, with reasoning, the most suitable group of sensitivity analysis methods for application to highly nonlinear dynamic models in the context of biological systems. Specifically, we perform computational experiments on antibody-producing mammalian cell culture models of different complexities and identify, as well as address, problems associated with such “real life” models. In conclusion, a novel global screening method (derivative based global sensitivity measures, DGSM) is proven to be the most time-efficient and robust alternative to the established variance-based Monte Carlo methods.

Polymer scaffolds tailored for tissue engineering applications possessing the desired pore structure require reproducible fabrication techniques. Nondestructive, quantitative methods for pore characterization are required to determine the pore size and its distribution. In this study, a promising alternative to traditional pore size characterization techniques is presented. We introduce a quantitative, nondestructive and inexpensive method to determine the pore size distribution of large soft porous solids based on the on the displacement of a liquid, that spreads without limits though a porous medium, by nitrogen. The capillary pressure is measured and related to the pore sizes as well as the pore size distribution of the narrowest bottlenecks of the largest interconnected pores in a porous medium. The measured pore diameters correspond to the narrowest bottleneck of the largest pores connecting the bottom with the top surface of a given porous solid. The applicability and reproducibility of the breakthrough technique is demonstrated on two polyurethane foams, manufactured using the thermally induced phase separation (TIPS) process, with almost identical overall porosity (60-70%) but very different pore morphology. By selecting different quenching temperatures to induce polymer phase separation, the pore structure could be regulated while maintaining the overall porosity. Depending on the quenching temperature, the foams exhibited either longitudinally oriented tubular macropores interconnected with micropores or independent macropores connected to adjacent pores via openings in the pore walls. The pore size and its distribution obtained by the breakthrough test were in excellent agreement to conventional characterization techniques, such as scanning electron microscopy combined with image analysis, BET technique, and mercury intrusion porosimetry. This technique is suitable for the characterization of the micro- and macropore structure of soft porous solids intended for tissue engineering applications. The method is sensitive for the smallest bottlenecks of the largest continuous pores throughout the scaffold that contributes to fluid flow.

The success of high-performance differential gel electrophoresis using fluorescent dyes (DIGE) depends on the quality of the digital image captured after electrophoresis, the DIGE enabled image analysis software tool chosen for highlighting the differences, and the statistical analysis. This study compares three commonly available DIGE enabled software packages for the first time: DeCyder V6.5 (GE-Healthcare), Progenesis SameSpots V3.0 (Nonlinear Dynamics), and Dymension 3 (Syngene). DIGE gel images of cell culture media samples conditioned by HepG2 and END2 cell lines were used to evaluate the software packages both quantitatively and subjectively considering ease of use with minimal user intervention. Consistency of spot matching across the three software packages was compared, focusing on the top fifty spots ranked statistically by each package. In summary, Progenesis SameSpots outperformed the other two software packages in matching accuracy, possibly being benefited by its new approach: that is, identical spot outline across all the gels. Interestingly, the statistical analysis of the software packages was not consistent on account of differences in workflow, algorithms, and default settings. Results obtained for protein fold changes were substantially different in each package, which indicates that in spite of using internal standards, quantification is software dependent. A future research goal must be to reduce or eliminate user controlled settings, either by automatic sample-to-sample optimization by intelligent software, or by alternative parameter-free segmentation methods.

Biotreatment technologies offer a cost-effective and efficient method for dealing with point-source releases of solvents. However, a major problem hampering these technologies is the fluctuating pollutant loads, which is especially critical for inhibitory pollutants. Provision of biotreatment systems able to cope with this problem is a significant technological and environmental challenge. This study investigates the potential for an absorber to act as buffer for shock loadings of inhibitory pollutants in waste-gas streams undergoing biological treatment. 1,2-Dichloroethane (DCE) was used as an example of a toxic and inhibitory organic pollutant. The stability of a combined oil-absorber-bioscrubber (OAB) system was compared to that of a bioscrubber only (BO) system when each was subjected to shock loads of DCE. The BO system was inoculated with Xanthobacter autotrophicus strain GJ10 and was submitted to sharp, sequential pulses in DCE inlet load, which caused system instability. Complete inhibition of the BO process occurred for a 3 h DCE pulse, leading to 9125 g of DCE m(-3)bioscrubber total organic discharged (TODDCE). Following the pulse, fluorescence in situ hybridization (FISH) showed that the active strain GJ10 was effectively washed-out. In contrast, the performance of the OAB system was stable during DCE shock loads. The carbon dioxide production remained stable, and low levels of effluent DCE and total organic carbon concentrations were found. For the 3 h pulse TODDCE was only 173 g of DCE m(-3)bioscrubber, and FISH indicated that the GJ10 strain remained active. We conclude that the OAB system offers an effective solution to the biological treatment of waste-gas containing fluctuating pollutant concentrations.