Holotomography-Based, Label-Free Quantification of Cellular Dry Mass as a Biophysical Indicator of Microglial Aβ Phagocytosis during Senescence

Senescent microglia undergo significant molecular and biochemical changes associated with impaired phagocytosis of amyloid-β (Aβ), a process implicated in neurodegenerative disease progression. However, quantitative biophysical metrics capable of capturing this functional impairment—and thus elucidating the role of microglial dysfunction in disease progression—remain limited. In this study, we identify cellular dry mass, measured by label-free holotomography, in combination with cell and nuclear morphological features, as sensitive biophysical indicators of microglial senescence and phagocytic capacity. Microglial senescence was induced using optimized hydrogen peroxide (H ₂ O ₂ ) treatment and validated through p21 and pRPS6 expression, and Raman spectroscopic signatures. Subsequently, phagocytosis assays were conducted following Aβ treatment. Results showed that dry mass showed a strong correlation with phagocytic decline in senescent cells. Senescent microglia treated with Aβ exhibited significantly lower dry mass and smaller cell size, but larger nuclear size, compared with Aβ-treated control microglia. These findings highlight dry mass as a robust, non-invasive biophysical indicator of microglial senescence and associated phagocytic function.