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Gewählte Publikation:

Walther, C; Kellner, M; Berkemeyer, M; Brocard, C; Durauer, A.
(2017): A microscale bacterial cell disruption technique as first step for automated and miniaturized process development
PROCESS BIOCHEM. 2017; 59: 207-215. FullText FullText_BOKU

Abstract:
Cell disruption is crucial during recovery of biopharmaceuticals overexpressed in E. coli, which tend to be produced intracellularly as insoluble inclusion bodies. Miniaturized high-throughput systems can accelerate the laborious downstream protocol for such biopharmaceuticals and enable integrated process-development. A fast and robust cell disruption method reflecting the protein and impurity profile of homogenates obtained by large-scale methods is required for such an approach. We established a miniaturized bead mill for parallel mechanical cell disruption at the microscale. Its total protein and impurity release, protein pattern, and particle size distribution were compared to results from microscale enzymatic digestion and referred to laboratory-scale high-pressure homogenization. Bead mill disruption led to equivalent protein and impurity release as well as to the same particle size profile as the large-scale reference. In contrast, lysates obtained by enzymatic digestion contained only 30-47% of overall protein, 17% of dsDNA, and 7-10% of endotoxin compared to those obtained by high-pressure homogenization; also larger debris was present in lysates after enzymatic digestion. The established method is fast, efficient, robust and comparable to current large-scale standards, allowing for parallelization of experiments. Thus, it is the method of choice for rapid integrated process development at the microscale.
Autor*innen der BOKU Wien:
Dürauer Astrid
Walther Cornelia
BOKU Gendermonitor:


Find related publications in this database (Keywords)
Bacterial disintegration
High-pressure homogenization
Scale up
High throughput
Inclusion body
Bead mill


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