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    A computational approach to quantifying miscounting of radiation-induced double-strand break immunofluorescent foci

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    Authors
    Ingram, Samuel
    Warmenhoven, John W
    Henthorn, Nicholas
    Chadwick, Amy
    Santina, Elham
    McMahon, S. J.
    Schuemann, J.
    Kirkby, Norman
    Mackay, Ranald I
    Kirkby, Karen J
    Merchant, Michael J
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    Affiliation
    Division of Cancer Sciences, Faculty of Biology, Medicine and Health, The University of Manchester, Oxford Rd, Manchester, M13 9PL, UK
    Issue Date
    2022
    
    Metadata
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    Abstract
    Immunofluorescent tagging of DNA double-strand break (DSB) markers, such as γ-H2AX and other DSB repair proteins, are powerful tools in understanding biological consequences following irradiation. However, whilst the technique is widespread, there are many uncertainties related to its ability to resolve and reliably deduce the number of foci when counting using microscopy. We present a new tool for simulating radiation-induced foci in order to evaluate microscope performance within in silico immunofluorescent images. Simulations of the DSB distributions were generated using Monte Carlo track-structure simulation. For each DSB distribution, a corresponding DNA repair process was modelled and the un-repaired DSBs were recorded at several time points. Corresponding microscopy images for both a DSB and (γ-H2AX) fluorescent marker were generated and compared for different microscopes, radiation types and doses. Statistically significant differences in miscounting were found across most of the tested scenarios. These inconsistencies were propagated through to repair kinetics where there was a perceived change between radiation-types. These changes did not reflect the underlying repair rate and were caused by inconsistencies in foci counting. We conclude that these underlying uncertainties must be considered when analysing images of DNA damage markers to ensure differences observed are real and are not caused by non-systematic miscounting.
    Citation
    Ingram SP, Warmenhoven JW, Henthorn NT, Chadiwck AL, Santina EE, McMahon SJ, et al. A computational approach to quantifying miscounting of radiation-induced double-strand break immunofluorescent foci. Communications biology. 2022 Jul 14;5(1):700. PubMed PMID: 35835982. Pubmed Central PMCID: PMC9283546. Epub 2022/07/15. eng.
    Journal
    Communications Biology
    URI
    http://hdl.handle.net/10541/625429
    DOI
    10.1038/s42003-022-03585-5
    PubMed ID
    35835982
    Additional Links
    https://dx.doi.org/10.1038/s42003-022-03585-5
    Type
    Article
    Language
    en
    ae974a485f413a2113503eed53cd6c53
    10.1038/s42003-022-03585-5
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