title: High-throughput cell quantification assays for use in cell purification development - enabling technologies for cell production.
abstract: High-throughput screening (HTS) technology is gaining increasing importance in downstream process development of cell-based products. The development of such HTS-technologies, however, is highly dependent on the availability of robust, accurate, and sensitive high-throughput cell quantification methods. In this article, we compare state-of-the-art cell quantification methods with focus on their applicability in HTS-platforms for downstream processing of cell-based products. Sensitivity, dynamic range, and precision were evaluated for four methods that differ in their respective mechanism. In addition, we evaluated the performance of these methods over a range of buffer compositions, medium densities, and viscosities, representing conditions found in many downstream processing methods. We found that CellTiter-Glo™ and flow cytometry are excellent tools for high-throughput cell quantification. Both methods have broad working ranges (3-4 log) and performed well over a wide range of buffer compositions. In comparison, CyQuant® Direct and CellTracker™ had smaller working ranges and were more sensitive to changes in buffer composition. For fast and sensitive quantification of a single cell type, CellTiter-Glo™ performed best, while for more complex cell mixtures flow cytometry is the method of choice. Our analysis will facilitate the selection of the most suitable method for a specific application and provides a benchmark for future HTS development in downstream processing of cell-based products.
authors.affiliation: Karlsruhe Institute of Technology (KIT), Institute of Process Engineering in Life Science, Section IV: Biomolecular Separation Engineering (MAB), Karlsruhe, Germany.
authors.forename: Sarah
authors.initials: S
authors.lastname: Zimmermann
authors.affiliation: Karlsruhe Institute of Technology (KIT), Institute of Process Engineering in Life Science, Section IV: Biomolecular Separation Engineering (MAB), Karlsruhe, Germany.
authors.forename: Sarah
authors.initials: S
authors.lastname: Gretzinger
authors.affiliation: Karlsruhe Institute of Technology (KIT), Institute of Process Engineering in Life Science, Section IV: Biomolecular Separation Engineering (MAB), Karlsruhe, Germany.
authors.forename: Christian
authors.initials: C
authors.lastname: Scheeder
authors.affiliation: Karlsruhe Institute of Technology (KIT), Institute of Process Engineering in Life Science, Section IV: Biomolecular Separation Engineering (MAB), Karlsruhe, Germany.|DIARECT AG, Department of Quality Assurance and Quality Control, Freiburg, Germany.
authors.forename: Marie-Luise
authors.initials: ML
authors.lastname: Schwab
authors.affiliation: Karlsruhe Institute of Technology (KIT), Institute of Process Engineering in Life Science, Section IV: Biomolecular Separation Engineering (MAB), Karlsruhe, Germany.|Boehringer Ingelheim Pharma GmbH & Co. KG, Global Bioprocess & Pharmaceutical Development, Biberach, Germany.
authors.forename: Stefan A
authors.initials: SA
authors.lastname: Oelmeier
authors.affiliation: Karlsruhe Institute of Technology (KIT), Institute of Process Engineering in Life Science, Section IV: Biomolecular Separation Engineering (MAB), Karlsruhe, Germany.
authors.forename: Anna
authors.initials: A
authors.lastname: Osberghaus
authors.affiliation: Karlsruhe Institute of Technology (KIT), Institute for Biological Interfaces (IBG 5), Eggenstein-Leopoldshafen, Germany.
authors.forename: Eric
authors.initials: E
authors.lastname: Gottwald
authors.affiliation: Karlsruhe Institute of Technology (KIT), Institute of Process Engineering in Life Science, Section IV: Biomolecular Separation Engineering (MAB), Karlsruhe, Germany. juergen.hubbuch@kit.edu.
authors.forename: Jürgen
authors.initials: J
authors.lastname: Hubbuch
mesh_terms: D049109:Cell Proliferation; D002470:Cell Survival; D005434:Flow Cytometry / Q000379:methods*; D018922:HL-60 Cells; D057166:High-Throughput Screening Assays / Q000379:methods*; D006801:Humans
keywords: Cell quantification assays; Cell-based products; Downstream processing; High-throughput screening (HTS)