Single-step photobleaching of a GFP monomer adsorbed on the glass and an Lyn-FG molecule in the PM observed at 200 Hz. Typical time-dependent signal intensity changes of the fluorescence spots (observed in 15 × 15–pixel areas) of a recombinant GFP molecule sparsely adsorbed on the coverglass (left) and an Lyn-FG molecule in the HeLa cell PM (right). A large stepwise decrease in each panel represents a single-step photobleaching of the fluorescence spot (arrowhead), indicating that virtually every fluorescence spot represented a single molecule. The signal level before photobleaching is shown by a red dashed line, whereas the background signal intensity is shown by a horizontal red solid line.