CFAssay: statistical analysis of the colony formation assay
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METHODOLOGY
Open Access
CFAssay: statistical analysis of the colony formation assay Herbert Braselmann1,2* , Agata Michna1 , Julia Heß1,2 and Kristian Unger1,2
Abstract Background: Colony formation assay is the gold standard to determine cell reproductive death after treatment with ionizing radiation, applied for different cell lines or in combination with other treatment modalities. Associated linear-quadratic cell survival curves can be calculated with different methods. For easy code exchange and methodological standardisation among collaborating laboratories a software package CFAssay for R (R Core Team, R: A Language and Environment for Statistical Computing, 2014) was established to perform thorough statistical analysis of linear-quadratic cell survival curves after treatment with ionizing radiation and of two-way designs of experiments with chemical treatments only. Methods: CFAssay offers maximum likelihood and related methods by default and the least squares or weighted least squares method can be optionally chosen. A test for comparision of cell survival curves and an ANOVA test for experimental two-way designs are provided. Results: For the two presented examples estimated parameters do not differ much between maximum-likelihood and least squares. However the dispersion parameter of the quasi-likelihood method is much more sensitive for statistical variation in the data than the multiple R2 coefficient of determination from the least squares method. Conclusion: The dispersion parameter for goodness of fit and different plot functions in CFAssay help to evaluate experimental data quality. As open source software interlaboratory code sharing between users is facilitated. Availability: The package is available at http://www.bioconductor.org/packages/release/bioc/html/CFAssay.html. Keywords: Colony formation assay, Cell survival, Linear-quadratic model
Background Clonogenic assay or colony formation assay (CFA) is an in vitro cell survival assay based on the ability of single cells to grow into colonies [1]. It is the gold standard to determine cell reproductive death after treatment with ionizing radiation. Whereby the relationship between the radiation doses and the proportion of surviving colonies is usually described by parametric cell survival curves. These can be used for the characterisation of the radiation sensitivity of different tumour cell lines given a specific radiation type [2], or in combination with other treatment modalities, e.g. a therapeutic agent or radiation sensitizer [3]. *Correspondence: [email protected] 1 Research Unit Radiation Cytogenetics, Helmholtz Zentrum München, German Research Center for Environmental Health GmbH, Neuherberg, Germany 2 Clinical Cooperation Group ’Personalized Radiotherapy of Head and Neck Cancer’, Helmholtz Zentrum München, German Research Center for Environmental Health GmbH, Neuherberg, Germany
For the analysis of cell survival curves CFAssay uses the commonly used linear-quadratic model (LQ model) [1]. Apart from radiation the CFA is al
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