HiIDDD: A high-throughput imaging pipeline for the quantitative detection of DNA damage in primary human immune cells

DNA damage is a prominent biomarker for numerous diseases, including cancer and aging. Detection of DNA damage routinely relies on traditional microscopy or cytometric methods. However, these techniques are typically of limited throughput and are not ideally suited for large-scale longitudinal and population studies that require analysis of large sample sets. We have developed HiIDDD ( Hi gh-throughput Immune cell D NA D amage D etection), a robust, semiquantitative and single-cell assay that measures DNA damage by high-throughput imaging using the two major DNA damage markers 53BP1 and γ−H2AX. We demonstrate sensitive detection of DNA damage in a wide set of freshly isolated and cryopreserved primary human immune cells, including CD4 + and CD8 + T cells, B cells and monocytes with low inter-assay variability. As proof of principle, we demonstrate parallel batch processing of several immune cell types from multiple donors. We find common patterns of DNA damage in multiple immune cell types of donors of varying ages, suggesting that immune cell properties are specific to individuals. These results establish a novel high-throughput assay for the evaluation of DNA damage in large-scale studies.