In Situ Detection of Complex DNA Damage Using Microscopy: A Rough Road Ahead.

Abstract

Simple SummaryWe comparatively discuss all possible methodologies for the complex DNA damage in situ detection. Fluorescent microscopy (FM) is the universal approach utilized in every technique, thus we discuss several FM variants. As image colocalization analysis is involved in the majority of methods, the related coefficients are reported. We envision this work to be a reference for radiobiologists in order to select and orchestrate the appropriate experimental and analysis strategies for optimized DNA damage detection. Last but not least, in the long term to help the incorporation of DNA damage biomarkers in the clinic as useful diagnostic or prognostic indicators.Complexity of DNA damage is considered currently one if not the primary instigator of biological responses and determinant of short and long-term effects in organisms and their offspring. In this review, we focus on the detection of complex (clustered) DNA damage (CDD) induced for example by ionizing radiation (IR) and in some cases by high oxidative stress. We perform a short historical perspective in the field, emphasizing the microscopy-based techniques and methodologies for the detection of CDD at the cellular level. We extend this analysis on the pertaining methodology of surrogate protein markers of CDD (foci) colocalization and provide a unique synthesis of imaging parameters, software, and different types of microscopy used. Last but not least, we critically discuss the main advances and necessary future direction for the better detection of CDD, with important outcomes in biological and clinical setups.