Optical tweezers use a highly focused laser beam to trap and manipulate dielectric particles in three dimensions. The technique exploits the gradient force that draws particles toward the focal point of the beam, balanced against scattering forces that push particles along the beam axis. The resulting optical trap can hold particles ranging from tens of nanometers to tens of micrometers with forces up to hundreds of piconewtons.

Forces are measured by detecting the displacement of the trapped particle from the trap center using back-focal-plane interferometry. A position-sensitive detector records the deflection of the trapping laser or a separate probe beam. The trap stiffness is calibrated by analyzing the Brownian motion power spectrum or by applying known viscous drag forces. Displacement sensitivity reaches 0.1 nm, and force sensitivity reaches 0.1 pN.

Optical tweezers are used to study molecular motors such as kinesin, myosin, and dynein, which generate piconewton forces while moving along cytoskeletal filaments. By attaching a motor protein to a trapped bead and measuring its stepping motion, researchers determine step size, force generation, and ATP coupling efficiency.

In protein folding and nucleic acid folding studies, optical tweezers and atomic force microscopy apply tension to individual molecules. The force-extension curve reports the mechanical stability of folded domains, unfolding intermediates, and folding pathways. RNA polymerase transcription, ribosome translation, and DNA helicase unwinding are studied by applying force to the nucleic acid substrate.

Dual-trap optical tweezers (dumbbell configuration) use two traps to hold the ends of a single molecule, eliminating surface attachment artifacts. Temperature-controlled stages and microfluidic sample delivery enable rapid buffer exchange and controlled biochemical conditions.