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Non-fluidic bio-layer interferometry (BLI) has rapidly become a standard tool for monitoring almost all biomolecular interactions in a label-free, real-time and high-throughput manner. High-efficiency screening methods which measure the kinetics of liposomes with a variety of compounds require the immobilization of liposomes. In this work, a method is described for immobilizing liposomes for interaction studies, based on the biophysical principles of this biosensor platform. The immobilization approach includes the loading of DSPE-PEG (2000) -biotin containing sterically stabilized micelles (SSMs) which are restructured in a buffer change step, resulting in an accessible substrate for liposome immobilization. Liposomes in a concentration of 5mM of varying composition and fluidity were immobilized on the sensor surface by inserting the hydrophobic residues of the former loaded SSMs. This proof of principle was carried out using Cytochrome C as a membrane-interacting model protein. The binding of Cytochrome C to the immobilized liposomes was demonstrated, and the derived kinetic and affinity constants were similar to values given in the literature. In order to obtain a detailed understanding of this surface, and to show the integrity of the liposomes, confocal fluorescence microscopy was used. Images of immobilized liposomes containing calcein in the aqueous core indicated intact vesicles. A combination of this simple liposome immobilization approach, the possibility of automation on BLI systems with high throughput within an acceptable timescale and excellent reproducibility makes this assay suitable for basic research as well as for industrial and regulatory applications., (Copyright © 2017 Elsevier B.V. All rights reserved.)

A new electrophoretic technique for the qualitative and quantitative analyses of IgM isoforms and fragments has been developed. IgMs which are more complex than many other recombinantly expressed immunoglobulins are characterized by their high molecular weighted active forms and many additional isoforms and fragments in the molecular range between 25 and 1200kDa. To analyze the multimers, isoforms and fragments simultaneously a high-resolution method, which enables sufficient migration and separation is required. Furthermore, this method should be appropriate to analyze IgMs in crude culture supernatants as well as purified samples. Simple sample preparation avoiding unspecific protein loss has been established. Currently no standard method to analyze all of them accordingly is available. The IgM-SDS-PAGE investigated for this purpose includes all these aspects. The combination of simple sample preparation and the application of precast gels make this electrophoretic method suitable for research but also quality control. The selective quantification of the multimers and the relative isoform distribution were performed by sensitive Sypro Ruby staining obtaining reliable and reproducible data in clone screening and process development which has been demonstrated by recombinantly expressed IgMs with significantly different isoform pattern., (2010 Elsevier B.V. All rights reserved.)

The scale-up of hydrophobic interaction chromatography is described. Human recombinant superoxide dismutase was used as a model. The scale-up was performed by keeping the height to diameter (H/D) ratio of the column constant. The success of scale-up was evaluated by reversed-phase high-performance liquid chromatography of the eluted material. The wrong H/D ratio causes decreased resolution.