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• W3209608975 abstract "The clonogenic assay measures the capacity of single cells to form colonies in vitro. It is widely used to identify and quantify self-renewing mammalian cells derived from in vitro cultures as well as from ex vivo tissue preparations of different origins. Varying research questions and the heterogeneous growth requirements of individual cell model systems led to the development of several assay principles and formats that differ with regard to their conceptual setup, 2D or 3D culture conditions, optional cytotoxic treatments and subsequent mathematical analysis. The protocol presented here is based on the initial clonogenic assay protocol as developed by Puck and Marcus more than 60 years ago. It updates and extends the 2006 Nature Protocols article by Franken et al. It discusses different strategies and principles to analyze clonogenic growth in vitro and presents the clonogenic assay in a modular protocol framework enabling a diversity of formats and measures to optimize determination of clonogenic growth parameters. We put particular focus on the phenomenon of cellular cooperation and consideration of how this can affect the mathematical analysis of survival data. This protocol is applicable to any mammalian cell model system from which single-cell suspensions can be prepared and which contains at least a small fraction of cells with self-renewing capacity in vitro. Depending on the cell system used, the entire procedure takes ~2–10 weeks, with a total hands-on time of <20 h per biological replicate. This Protocol Extension discusses several approaches to analyzing clonogenic growth of mammalian cells in vitro, using a modular framework to facilitate the use of various formats to fully optimize clonogenic growth." @default.
• W3209608975 created "2021-11-08" @default.
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• W3209608975 date "2021-10-25" @default.
• W3209608975 modified "2023-10-17" @default.
• W3209608975 title "Analysis of clonogenic growth in vitro" @default.
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• W3209608975 doi "https://doi.org/10.1038/s41596-021-00615-0" @default.
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• W3209608975 hasPublicationYear "2021" @default.
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