A Comprehensive Framework for Spatio-Temporal Analysis of DNA Damage Foci in Tumor Spheroids.

The evaluation of DNA damage response, particularly DNA damage foci formation, is crucial for understanding tumor biology and assessing the impacts of various drugs. We have developed a sophisticated semi-automated image analysis pipeline which generates quantitative map of the spatiotemporal distribution of DNA damage foci within live tumor spheroids. Our framework seamlessly integrates live imaging of tumor spheroids via Light Sheet Fluorescence Microscopy with a DNA damage foci formation assay using a genetically encoded fluorescently labeled DNA damage sensor. By combining advanced imaging techniques with computational tools, our framework offers a powerful tool for studying DNA damage response mechanisms in complex 3D cellular environments.