Improved Axial Optical Trapping

Conventional optical tweezers suffer from several complications when applying axial forces to surface-tethered molecules. Aberrations from the refractive-index mismatch between an oil-immersion objectiveu0027s medium and the aqueous trapping environment degrade the trapping strength with focal depth. Furthermore, interference effects from back-scattered light make it difficult to use back-focal-plane interferometry for high-bandwidth position detection. Holographic optical tweezers were employed to correct for aberrations to achieve a constant axial stiffness and modulate artifacts from backscattered light. By discarding light from the centre of the trapping beam, interference from back-reflections could be reduced, but not eliminated without significant degradation to the trapping strength. However, careful calibration of the detector sensitivity allows for the application and measurement of axial forces on small surface-tethered molecules. The use of a donut beam to reduce the enhanced photobleaching of fluorophores by the trapping light was also investigated with the goal of combining our holographic axial tweezers with single-molecule fluorescence.