Improved Nanometer-Scale Particle Tracking in Optical Microscopy using Microfabricated Fiduciary Posts

Abstract

Pillars of Truth Small vibrations that occur during microscopy can often lead to big problems with the visualization of single molecules or the interpretation of subcellular processes. Often these small vibrations come from the microscope itself, caused by instrument parts such as a motorized stage or an attached CCD camera. This can become especially problematic when the experimental goal is to track the nanoscale movement of cellular organelles such as vesicles. In this issue, Dangaria et al. provide an inexpensive fix to this problem by microfabricating fiduciary posts attached to coverslips and using these posts to determine amounts of low frequency positional drift. The posts, 10 µm high by 3 µm in diameter, allow tracking of coverslip motion, thus providing a means to subtract coverslip motion or drift from the movement of cellular organelles. The authors examined the amount of drift due to vibrations from a motorized stage, finding an average drift velocity of 0.3 µm/s. This coverslip drift could t...