Figure 2

Tuning of STED depletion power. (a) Representative images of U937-PR9 cells upon staining of DNA replication foci through incorporation of EdU labeled with Alexa azide 488 (Click reaction). (top) Sequential acquisition of a confocal image, followed by a STED image with a 9 mW depletion beam. (bottom) Sequential acquisition of a confocal image, followed by a STED image with a 18 mW depletion beam. Scale bars represent 3 µm. (b) Line profiles of structures from images in (a). (top) Comparison between the line profiles of the same structure in the confocal (black line) and the STED (red line) images in the top row of (a). The measured structure is defined by a white dotted line in the confocal image. (bottom) Comparison between the line profiles of the same structure in the confocal (black line) and the STED (red line) images in the bottom row of (a). The measured structure is defined by a white dotted line in the confocal image. (c–e) Quantification of Resolution, Brightness, and Noise parameters by application of the QuICS algorithm. At least ten images for each condition have been acquired. Error bars represent the standard error of the mean (SEM) and ***p < 0.001.