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July 2nd, 2025
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Nanyang Technological University
bioengineering
biorxiv

Label-Free Mapping of Subcellular Dynamics using Wide-field Interferometric Scattering Microscopy and Spectral Exponent Analysis

Anyi, C. L.Open in Google Scholar•You, H.Open in Google Scholar•Li, H.Open in Google Scholar•Goh, K. L.Open in Google Scholar•Ling, T.Open in Google Scholar

Interferometric scattering (iSCAT) microscopy combines label-free detection with nanometer-scale motion sensitivity. It captures temporal signal fluctuations from sub-diffraction movements of scatterers in live cells, including macromolecules and cell membranes. Here, we investigate label-free mapping of subcellular dynamics by calculating the power spectral density (PSD) of the iSCAT time series. Using a wide-field iSCAT, we observe an inverse-power-law relationship S(f) = {beta}f- over 30 - 1250 Hz in multiple cell types. We compute pixel-wise PSDs and obtain a color-coded spectral exponent map based on the fitted spectral exponent () and amplitude ({beta}), which indicates the characteristics and strength of subcellular movements. We also incorporate the goodness-of-fit as color saturation to enhance the contrast of the spectral exponent map. Following this approach, we demonstrate label-free imaging of live cells while effectively distinguishing between benign and malignant thyroid cancer subtypes, mitotic and interphase cells, as well as live and apoptotic cells. Dynamic imaging using wide-field iSCAT provides an intrinsic, label-free marker of cellular states and mechanical properties, which may benefit studies in mechanobiology, cancer diagnostics, and stem cell therapies.

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