Abstract

Ionizing radiation induces amongst other the most critical type of DNA damage: double-strand breaks (DSBs). Efficient repair of such damage is crucial for cell survival and genomic stability. The analysis of DSB associated foci assays is often performed manually or with automatic systems. Manual evaluation is time consuming and subjective, while most automatic approaches are prone to changes in experimental conditions or to image artefacts. Here, we examined multiple machine learning models, namely a multi-layer perceptron classifier (MLP), linear support vector machine classifier (SVM), complement naive bayes classifier (cNB) and random forest classifier (RF), to correctly classify γH2AX foci in manually labeled images containing multiple types of artefacts. All models yielded reasonable agreements to the manual rating on the training images (Matthews correlation coefficient >0.4). Afterwards, the best performing models were applied on images obtained under different experimental conditions. Thereby, the MLP model produced the best results with an F1 Score >0.9. As a consequence, we have demonstrated that the used approach is sufficient to mimic manual counting and is robust against image artefacts and changes in experimental conditions.

Highlights

  • The effect of irradiation on bio-chemical properties of cells or their survival can be evaluated by detection of the induced DNA damage either in the form of single strand breaks, double strand breaks or others

  • The present study was performed to develop a method for automatic detection of γH2AX foci, using standard fluorescence microscopy, resulting in images of strongly varying quality

  • The used models were surprisingly robust against artefacts and changes in experimental conditions, potentially allowing a generalization of the best performing multi-layer perceptron classifier (MLP) model, after initial training

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Summary

Introduction

The effect of irradiation on bio-chemical properties of cells or their survival can be evaluated by detection of the induced DNA damage either in the form of single strand breaks, double strand breaks or others. Double strand breaks are considered to be the most toxic form of DNA damage [1] as these lesions are significantly more complex in nature and time-consuming to repair [2,3]. A common way to visualize DNA damage or more precisely double strand breaks is via labeling of γH2AX the phosphorylated form of the H2AX histone protein variant. ΓH2AX is a sensitive marker for DNA damage, as the number of γH2AX foci is proportional to the dose of irradiation [6,7,8,9]. H2AX was shown to be part of the DNA damage signaling, located closely to DNA doublestrand breaks [4,5]. γH2AX is a sensitive marker for DNA damage, as the number of γH2AX foci is proportional to the dose of irradiation [6,7,8,9].

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