Kinetics from single-molecule fluorescence trajectories and single-photon time series. (a) A representation of a ribosome structure with different structural features and labelling sites is shown on the left. On the right, a single-molecule fluorescence trajectory (green, donor; red, acceptor) and resulting FRET trajectory (blue) obtained using a wide-field TIRF imaging system at 40 ms time resolution under equilibrium conditions from a surface-immobilized ribosome bearing site-specifically labelled A-site and P-site tRNA molecules . The dynamic FRET data, reporting directly on thermally accessible, nanometre-scale changes in tRNA position within the ribosome, persist until donor and/or acceptor fluorophore photobleach. Figure from ref. . (b) On the left, a photon time series (green, donor; red, acceptor) from a confocal measurement using single-photon counting is shown. A state trajectory obtained from the Viterbi algorithm is shown as a blue line, with segments corresponding to folded and unfolded protein, and transition points . On the right, a cartoon of the structure of the small protein α3D immobilized on a surface is shown, whose folding was investigated here. Figure adapted from ref. .