Untersuchungen von Biomaterial - Bead Wechselwirkungen in der Expanded Bed Adsorption durch AFM Kraft-Distanz Messungen und DLVO Theorien
Zusammenfassung der Projektergebnisse
Expanded Bed Adsorption (EBA) is a bioprocessing technique that helps in the purification of products which are manufactured by cell cultures for therapeutic or diagnostic use in a cost effective and environmentally sustainable way. It is also one of the main technologies targeted within the large scale European project INTENSO. Improving the understanding and performance of EBA could have a profound impact on the development of new drugs, the availability of innovative medical solutions, and the reduction of the costs associated with such novel therapeutic schemes. This DFG project has advanced our understanding of some of the basic interactions taking place in EBA: the interaction or adhesion between cells and small chromatographic beads. Unfavorable adhesion of cells to beads can degrade the efficiency and the process performance of EBA. In this project the cell-bead interactions were investigated and characterized by two complementary methods: Global parameters like charge and contact angle of the materials were used as input for XDLVO theory to describe the adhesion of cells to beads. Here we improved the technology of the measurements by simplifying and increasing the range of contact angle and zeta potential measurements. Most chromatography users tend to ignore the effects that the bead matrix might have on product recovery or chromatographic performance. We collected a large data set for different cells, bead materials, and process parameters, and were able to extract some general relation between bead properties and the adhesion of cells such as bacteria, yeast, or mammalian cells. We particularly targeted conditions that are close to situations from practice and that represent current bottlenecks, e.g. the purification of therapeutic antibodies from high density cell cultures. The results can help to optimize the process parameters used in EBA. By a second approach we did not calculate but measured directly the forces between individual beads and cells used in EBA. This required a sensitive method - atomic force microscopy - which comes with a more tedious experimental setup and procedure: small changes or variations in experimental parameters can significantly influence the results. But with enough statistics our results confirmed for the first time the general use of XDLVO theory to describe interactions taking place in EBA. Our results also showed some of the limits of the theoretical description for a specific process situation, e.g. when hydrophobic interactions are the driving factor for product capture. This project allowed us to establish multiple connections with the private industry and to further test the validity of our methods under real operating conditions. As an example, our methods helped to improve the yield of recovery and the level of purity of a molecule. The methods developed, and the studies pursued during this project have helped us to understand - for the first time - the extent and nature of mammalian cell deposition and to improve mammalian cell-based bioprocessing systems. The project has generated methods that could guide work in the biopharmaceutical industries to produce in a more efficient manner improved chromatographic beads and biological agents that are used to attack difficult-to-treat illnesses.
Projektbezogene Publikationen (Auswahl)
- Extended DLVO calculations expose the role of the structural nature of the adsorbent beads during chromatography, Journal of Separation Science 35 (2012) 1068-1078
Aasim, Muhammad; Bibi, Noor Shad; Vennapusa, Rami Reddy; et al.
(Siehe online unter https://doi.org/10.1002/jssc.201100719) - Utilization of surface energetics approach to understand protein interaction to ceramic hydroxyapatite, Journal of Chemical Technology and Biotechnology 88 (2013) 1421-1428
Aasim, Muhammad; Bibi, Noor Shad; Vennapusa, Rami Reddy; et al.
(Siehe online unter https://doi.org/10.1002/jctb.4065) - The role of ligands on protein retention in adsorption chromatography: A surface energetics approach, Journal of Separation Science 37 (2014) 618-624
Aasim, Muhammad; Kakarla, Prasad Babu; D'Souza, Roy N.; et al.
(Siehe online unter https://doi.org/10.1002/jssc.201301338) - Growth-dependent Surface Characteristics of Hansenula polymorpha: Implications for Expanded Bed Adsorption Chromatography, Biotechnology and Bioprocess Engineering 20 (2015) 576-584
Naz, Nadia; Dsouza, Roy N.; Yelemane, Vikas; et al.
(Siehe online unter https://doi.org/10.1007/s12257-014-0397-6) - Interactions of Chinese Hamster Ovary (CHO) cell cultures with second generation expanded bed adsorbents, Separation and Purification Technolog 144 (2015) 23-30
Kakarla, Prasad Babu; DSouza, Roy N.; Toots, Urve; et al.
(Siehe online unter https://doi.org/10.1016/j.seppur.2015.02.014)
