Fig. 4

Study of cellular uptake and intracellular localization of PDA-ICG nanoprobe on SK-MEL-103 and A549 cells using confocal fluorescence microscopy and flow cytometry. a. Schematic illustration showing the cellular uptake of PDA-ICG by either cancer cells or senescent cells. Cells were first validated for senescence 10 days following treatments with either chemotherapeutic drugs or vehicles. b. Representative images of b-galactosidase staining of fixed control (vehicle-treated) cells, palbociclib-treated SK-MEL-103 cells and cisplatin-treated A549 cells. c. Western blot analysis of the expression of relevant senescence markers: phosphorylated retinoblastoma (pRb), and cell cycle inhibitor p21 with β-actin as reference. d. Confocal microcopy images of cells following 24-h incubation with 10 µg/ml PDA-ICG nanoparticles. Cell membranes, nucleus and lysosomes were stained with CellMask (green), Hoechst 33,342 (blue) and LysoTracker (yellow). PDA-ICG was observed in the Alexa Fluor 700 channel (red). Scale bar = 20 μm. e. Corresponding fluorescence intensity for nanoprobe-treated cells, showing senescent cells having higher PDA-ICG fluorescence intensity compared to cancer cells (mean ± SD, n = 3, ** for p ≤ 0.01, **** for p < 0.0001). f. Histogram showing mean fluorescence values of SK-MEL-103 cells after incubation with PDA-ICG; PDA-ICG fluorescence was measured from the Alexa Fluor 700 channel that corresponded to the fluorescence of PDA-ICG nanoparticles.