Automated high-throughput methods that support tracking of mammalian cell growth are currently needed to advance cell line characterization and identification of desired genetic components required for cell engineering. Here, we describe a high-throughput noninvasive assay based on plate reader measurements. The assay relies on the change in absorbance of the pH indicator phenol red. We show that its basic and acidic absorbance profiles can be converted into a cell growth index consistent with cell count profiles, and that, by adopting a computational pipeline and calibration measurements, it is possible to identify a conversion that enables prediction of cell numbers from plate measurements alone. The assay is suitable for growth characterization of both suspension and adherent cell lines when these are grown under different environmental conditions and treated with chemotherapeutic drugs. The method also supports characterization of stably engineered cell lines and identification of desired promoters based on fluorescence output.

Simultaneous Plate-Reader Characterization of Promoter Activity and Cell Growth in Engineered Mammalian Cells / Grob, Alice; Enrico Bena, Chiara; Redwood-Sawyerr, Chileab; Polizzi, Karen; Bosia, Carla; Isalan, Mark; Ceroni, Francesca (METHODS IN MOLECULAR BIOLOGY). - In: Synthetic Promoters : Methods and Protocols / Marchisio M.A.. - [s.l] : Humana Press - Springer, 2024. - ISBN 9781071640623. - pp. 85-96 [10.1007/978-1-0716-4063-0_5]

Simultaneous Plate-Reader Characterization of Promoter Activity and Cell Growth in Engineered Mammalian Cells

Enrico Bena, Chiara;Bosia, Carla;
2024

Abstract

Automated high-throughput methods that support tracking of mammalian cell growth are currently needed to advance cell line characterization and identification of desired genetic components required for cell engineering. Here, we describe a high-throughput noninvasive assay based on plate reader measurements. The assay relies on the change in absorbance of the pH indicator phenol red. We show that its basic and acidic absorbance profiles can be converted into a cell growth index consistent with cell count profiles, and that, by adopting a computational pipeline and calibration measurements, it is possible to identify a conversion that enables prediction of cell numbers from plate measurements alone. The assay is suitable for growth characterization of both suspension and adherent cell lines when these are grown under different environmental conditions and treated with chemotherapeutic drugs. The method also supports characterization of stably engineered cell lines and identification of desired promoters based on fluorescence output.
2024
9781071640623
9781071640630
Synthetic Promoters : Methods and Protocols
File in questo prodotto:
File Dimensione Formato  
2024_MethodsInMolecularBiology_Ceroni-Capitolo_5.pdf

accesso riservato

Tipologia: 2a Post-print versione editoriale / Version of Record
Licenza: Non Pubblico - Accesso privato/ristretto
Dimensione 3.35 MB
Formato Adobe PDF
3.35 MB Adobe PDF   Visualizza/Apri   Richiedi una copia
Pubblicazioni consigliate

I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.

Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11583/2996419