Label-free classification of growth-arrested state of cells by quantitative phase microscopy in fixative-treated cells

In the recurrence of cancer and autoimmune diseases, cellular senescence, a state where cells cease division, plays a pivotal role. Distinguishing the senescence requires a combination of multiple markers. Fluorescent dyes incorporation limits the imaging observation time due to the potential toxicity of those compounds over prolonged exposure, while colorimetric dye incorporation requires cell fixation, where cells are not in native state, and long staining time (≥4 hours). One of the distinct characteristics that distinguishes senescent cells from other state of cells is the impaired protein degradation ability by the lysosome, resulting on the protein accumulation. In the current work, we take advantage of protein accumulation to distinguish senescence cells from other cells. The protein concentration is visualized by recording the phase difference using the quantitative phase microscopy (QPM). Our live QPM observations on breast cancer cells confirm that senescent cells exhibit a higher refractive index compared to cells in normal growth and quiescent states, attributed to the accumulation of undigested lysosomal protein cargo. This heightened refractive index suggests an imbalance in protein turnover rates within senescent cells. These findings shed light on a potential label-free non-invasive approach for a prolonged monitoring of senescent cell dynamics.