Researchers at Nanyang Technological University, Singapore, have developed a label-free imaging strategy that maps subcellular dynamics in living cells using wide-field interferometric scattering (iSCAT) microscopy, an optical technique with nanometer-scale sensitivity. The study, published in PhotoniX Life, shows that statistical analysis of stochastic fluctuations in iSCAT image sequences can reveal cellular states and dynamic heterogeneity without added labels.
Cells are dynamic systems. Internal organelles, macromolecules, membranes, and cytoskeletal structures undergo thermal and active motions that are associated with both subcellular architectures and energy states. However, directly visualizing these fast, spatially heterogeneous motions in living cells remains challenging, and they may also be perturbed by exogenous labels commonly used in biological studies.
To address this challenge, the team performed a full-field analysis of the power spectral density (PSD) of high-speed iSCAT time series. Across multiple cell types, they observed that the PSD of iSCAT signals from most cellular regions followed an inverse-power-law relationship, Sf=βf-α, over 30–1,250 Hz. The fitted spectral exponent (α) and amplitude (β) reflect the characteristics and strength of subcellular movement. By encoding α as hue, β as value, and goodness-of-fit (R2) as saturation in the hue-saturation-value (HSV) space, they generated two-dimensional spectral exponent maps that reflect the spatial distribution of cellular dynamics.
Using this approach, the researchers demonstrated label-free visualization of cell-state changes. In HeLa cells, spectral exponent maps highlighted differences between mitotic and interphase cells and tracked dynamic transitions during mitotic progression. In hydrogen peroxide-induced apoptotic cells, the maps revealed increased spectral exponent heterogeneity and regions with high α associated with apoptosis-related membrane blebbing. The maps also captured malignancy-associated differences among thyroid cancer cell subtypes: papillary thyroid carcinoma (PTC), follicular thyroid carcinoma (FTC), and anaplastic thyroid carcinoma (ATC) cells showed progressively lower median spectral exponent values with increasing malignancy.
These results suggest that wide-field iSCAT imaging combined with power spectral exponent analysis may serve as an intrinsic optical readout of cellular states. Because the method does not require exogenous labels, it may be particularly useful for longitudinal live-cell studies, mechanobiology, cancer research, and quality assessment in stem-cell therapies.
See the article:
"Label-free mapping of subcellular dynamics using wide-field interferometric scattering microscopy and spectral exponent analysis." PhotoniX Life. doi: 10.3724/PXLIFE.2025-0012
https://doi.org/10.3724/PXLIFE.2025-0012
