Atomic Force Microscopy for Cellular Stiffness

Cells were plated at the density of 3 × 10³ cells/cm², and cellular stiffness on top of the nucleus and the cytosol was measured using atomic force microscopy (AFM) indentation method 29 (link)–31 (link) by the JPK NanoWizard II AFM system (JPK Instruments, Berlin, Germany). The cantilever was the tipless cantilever (ARROW-TL1-50; NanoWorld, Neuchatel, Switezerland), which conjugated with a polystyrene bead (with diameter of 5 μm). Before each measurement, cantilever was calibrated and possessed a mean spring constants ranging from 0.01 to 0.03 nN/nm. For stiffness measurements, 1 nN indentation was applied to each cell. The indentation depth was about 0.5 μm to avoid the substratum effect. The approaching speed of the cantilever to cell surface was set at 1 μm/sec. Force-distance curves were collected by landing the bead on region of interests and processed using JPK IP software. To extract the mechanical properties of cells from force-distance curves, the data were analysed by the Hertz model.