Multiplexed Chromatin Analysis Using Optical Spectroscopic Statistical Nanosensing.

In single cells, chromatin packs into organized structures to perform biological functions, such as RNA transcription regulation. Characterizing such structural behaviors, including packing density and mass scaling, is critical in epigenetics research. Partial wave spectroscopic (PWS) microscopy is a label-free, live-cell, high-throughput imaging modality that utilizes optical spectroscopic statistical nanosensing. Rather than resolving the exact chromatin packing structure, PWS extracts statistical packing information from spectroscopic interference signals. In this study, we evaluate its ability to characterize multiplexed chromatin packing density and mass scaling, as well as its spatial confidence interval, using finite difference time domain (FDTD) electromagnetic simulations. We validated the simulation-based analysis algorithm by comparing experimental PWS images against coregistered super-resolution acquisitions, confirming its accuracy in capturing chromatin packing metrics. We then applied this modality to live cells treated with different epigenetic agents, mapping spatial changes in chromatin packing in a high-throughput workflow.