Near-physiological cell cycle synchronization with countercurrent centrifugal elutriation

Journal
Methods in molecular biology  
Start Page
3
End Page
16
Citation
Methods in Molecular Biology (2095): 3-16 (2020)
Publisher DOI
10.1007/978-1-0716-0191-4_1
Scopus ID
2-s2.0-85077143916
The bioreactor conditions and cell diversity in mammalian cell cultures are often regarded as homogeneous. Recently, the influence of various kinds of heterogeneities on production rates receives increasing attention. Besides spatial gradients within the cultivation system, the variation between cell populations and the progress of the cells through the cell cycle can affect the dynamics of the cultivation process. Strong metabolic up- and down-regulations leading to variable productivities, even in exponentially growing cell cultures, have been identified in CHO cell cultivations. Consequently, scientific studies of cell cycle-related effects and metabolic regulations require experiments utilizing cell cycle-enriched subpopulations. Importantly, the enrichment procedure itself must not strongly interfere with the cell culture under investigation. Such subpopulations can be generated by near-physiological countercurrent centrifugal elutriation, which is described in the following chapter. At first, a brief overview regarding the cell cycle, currently identified effects and commonly used methods, and their applicability is outlined. Then, the experimental setup and the synchronization itself are explained.
Subjects
Cell size
Centrifugation
Population heterogeneities
Separation