Table 1.

Summary of pros and cons of the various combined visualization and manipulation techniques described

Technique	Pros	Cons
Molecular Combing	Technically simple Many DNA molecules can be studied in parallel TIRF compatible Compatible with low protein concentrations (∼10 pM)	Multiple uncontrolled attachment points No control of force Slow buffer exchange
Surface-tethered DNA	Many DNA molecules can be studied in parallel TIRF compatible Single attachment point to surface Can be compatible with low protein concentrations (∼10 pM)	Little force control Slow buffer exchange Requires continuous smooth fluid flows
DNA fixed to one optically trapped bead	Fast buffer exchange Some force control No background fluorescence from surface	Requires continuous smooth fluid flows Minimum protein concentration ∼1 nM because of microfluidics system (46) Technically more challenging
DNA tethered between two optically trapped beads	Fast buffer exchange High degree of force control Direct measurement of force on DNA No background fluorescence from surface (using water immersion objective)	Technically challenging Minimum protein concentration ∼1 nM because of microfluidics system (46)