DNase activity in invadosomes reported by SNS. (a) Basic invadosome structure and representative proteins. DNase (depicted in green) is revealed in this study. (b) Surface-immobilized Cy3-labeled dsDNA visualizes DNase activity by fluorescence loss. (c) SNS with a quencher–dye pair reports DNase activity by fluorescence gain. (d) A THP-1 cell on a surface coated with Cy3-labeled dsDNA. Podosomes were identified by the characteristic actin core and vinculin ring. dsDNA was degraded in a dark punctate pattern colocalized with the podosomes. BF, bright field. (e) A THP-1 cell on an SNS-immobilized surface, which showed DNA degradation in a bright punctate pattern colocalized with podosomes. Statistics show that 86 ± 5% of actin puncta are colocalized with SNS punctate signal (600 podosomes over 20 cells). (f and g) Representative regions marked by yellow boxes in panels d and e, respectively. Line profiles (marked by yellow lines) of vinculin, F-actin, and DNase sensor signal in single podosomes are also presented in panels f and g. Scale bars represent 5 µm (d) and 1 µm (f and g).