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The fluorometric microculture cytotoxicity assay

Nature Protocols volume 3, pages 1364–1369 (2008)Cite this article

Abstract

The fluorometric microculture cytotoxicity assay (FMCA) is a nonclonogenic microplate-based cell viability assay used for measurement of the cytotoxic and/or cytostatic effect of different compounds in vitro. The assay is based on hydrolysis of the probe, fluorescein diacetate (FDA) by esterases in cells with intact plasma membranes. The assay is available as both a semiautomated 96-well plate setup and a 384-well plate version fully adaptable to robotics. Experimental plates are prepared with a small amount of drug solution and can be stored frozen. Cells are seeded on the plates and cell viability is evaluated after 72 h. The protocol described here is applicable both for cell lines and freshly prepared tumor cells from patients and is suitable both for screening in drug development and as a basis for a predictive test for individualization of anticancer drug therapy.

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Figure 1: Relationship between cell density and signal-to-noise ratio in the fluorometric microculture cytotoxicity assay (FMCA).
Figure 2: Standard plate layout of a 384-well microtiter plate for dose–response characterization of 18 drugs.
Figure 3: Reproducibility of the effect of four drugs on the myeloma cell line, 8226/S, on different plate batches and assay time-points during 1 year.

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Authors and Affiliations

  1. Division of Clinical Pharmacology, Department of Medical Sciences, Uppsala University, Uppsala University Hospital, Uppsala, SE-751 85, Sweden

    Elin Lindhagen & Rolf Larsson

  2. Division of Oncology, Department of Oncology, Radiology and Clinical Immunology, Uppsala University, Uppsala University Hospital, Uppsala, SE-751 85, Sweden

    Peter Nygren

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  1. Elin Lindhagen
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Correspondence to Elin Lindhagen.

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Lindhagen, E., Nygren, P. & Larsson, R. The fluorometric microculture cytotoxicity assay. Nat Protoc 3, 1364–1369 (2008). https://doi.org/10.1038/nprot.2008.114

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