Figure 4
Simultaneous SR-SIM/AFM acquisition. The AFM measurements were carried out on fixed U2OS cells in medium/buffer with (a) and without N-SIM illumination (b). For convenience and enhanced feature/noise contrast, both AFM topography images in the SR-SIM/AFM overlays are displayed with an edge detection algorithm using a pixel difference operator in X. The topography images from Petri dish surface on three positions (labelled in the figures) were planefit (1st order polynomial function) to compensate for tilts in the sample surface, and subjected to surface roughness analysis (c). For comparison reasons, the average roughness (Ra), RMS roughness (Rq) and peak-to-valley roughness (Rt) values are given below the corresponding height profiles. XY-scales of the AFM images in (a,b) are 13.59 μm and 12.16 μm respectively, recorded at a resolution of 256 × 229 pixels. The insets used for analysis in (c) have a resolution of 37 × 37 pixels. SR-SIM images were obtained using NIS-Elements (v5.11.00), while the overlay of the SR-SIM and AFM data were generated with the JPK Data Processing software (v6.1.120).
