Utilization of Holotomographic and Live-Cell Fluorescent Microscopy to Study the U2OS Cell Cycle

Cancer is a leading cause of death worldwide, causing 1 in 6 deaths globally [7]. A hallmark of cancer progression is cell cycle dysregulation, with proto-oncogene mutations and the loss of tumor suppressor genes causing accelerated, uncontrolled cell division. Currently, fluorescence microscopy serves as a tool to observe cell cycle progression and to localize cell-cycle-dependent proteins. However, fluorescence microscopy introduces unwanted artifacts, such as photobleaching and phototoxicity, resulting in an inability to observe long-term dynamic changes to cells. Holotomographic microscopy (HTM) is an emerging technique that measures refractive index (RI) signatures, granting real-time, label-free visualization of cells and their subcellular components. This non-destructive and non-interfering method offers unprecedented opportunities to study cell cycle progression using RI signatures. Here, we aim to integrate HTM imaging with fluorescence microscopy to investigate correlations between RI signatures of the nucleus and different stages of the cell cycle. By correlating RI signatures to cell cycle progression, this approach can develop a label-free marker of each cell cycle stage, granting label-free, non-destructive imaging of the cell cycle.