Chapter 1. Direct fluorescent imaging of translocation and unwinding by individual DNA helicases -- Chapter 2. Single-molecule imaging with one color fluorescence -- Chapter 3. Measuring membrane protein dimerization equilibrium in lipid bilayers by single-molecule fluorescence microsopy -- Chapter 4. Fluorescent labeling of proteins in whole cell extracts for single-molecule imaging -- Chapter 5. Quantifying the assembly of multicomponent molecular machines by single-molecule total internal reflection fluorescence microscopy -- Chapter 6. How to measure separations and angles between intramolecular fluorescent markers -- Chapter 7. Precisely and accurately inferring single-molecule rate constants -- Chapter 8. Quantification of functional dynamics of membrane proteins reconstituted in nanodiscs membranes by single turnover functional readout -- Chapter 9. Putting Humpty-Dumpty together: clustering the functional dynamics of single biomolecular machines such as the splicosome -- Chapter 10. Single-molecule FRET to measure conformational dynamics of DNA mismatch repair proteins -- Chapter 11. Single-molecule confocal FRET microscopy to dissect conformational changes in the catalytic cycle of DNA topoisomerases -- Chapter 12. Probing the conformational landscape of DNA polymerases using diffusion-based single-molecule FRET -- Chapter 13. Methods for investigating DNA accessibility with single nucleosomes -- Chapter 14. Single-molecule fluorescence studies of fast protein folding -- Chapter 15. Single-molecule multicolor FRET assay for studying structural dynamics of biomolecules -- Chapter 16. A multicolor single-molecule FRET approach to study protein dynamics and interactions simultaneously -- Chapter 17. Interferometric scattering microscopy for the study of molecular motors -- Chapter 18. Enzyme kinetics in femtoliter arrays -- Author index -- Subject index.