Combining confocal microscopy with precise force-scope optical tweezers

Abstract

We demonstrate an example of 'confocal-tweezers' wherein confocal images and precise optical force measurements, using photodiodes, are obtained simultaneously in the x-y plane without moving the objective lens. The optical trap is produced using a 1.064&#956;m cw laser and is combined with Leica's TCS SP5 broadband confocal microscope to trap and image living cells. The unique method by which the confocal images are created facilitates the acquisition of images in areas far from the trapping location. In addition, because the scanning process involves moving galvanic mirrors independently of the objective, the trap is held stable in position and is not subject to any error in position for the x-y scan. We have successfully trapped and confocally imaged 80nm gold colloids, 150nm gold colloids and 1&#956;m polystyrene beads whilst making quantitative measurements of the force applied by the trap on each bead. To the best of our knowledge this is the first time that anyone has combined precise force measuring optical tweezers with confocal microscopy. We also discuss some of the technical challenges involved in advancing the experimental set up to make quantitative force measurements in combination with 3D stacking. Having proven the potential of this system in 2D, we hope to develop it further to investigate the nano-mechanics of cell division through the attachment of gold beads to fluorescently labelled organelles in <i>S. pombe </i>yeast cells.