In Vivo Organelle Tracking, Calibration, and Force Measurement with an Optical Trap

Abstract

We report our results on calibrating an optical trap in vivo while simultaneously trapping and tracking organelles with it. We have built an optical trap with microsecond time resolution and nm spatial resolution with a trapping laser that is positioned with an Acousto-Optic Deflector (AOD). This AOD is of fundamental importance to the in vivo stiffness calibration technique, as this technique requires high speed oscillation of the trap position (on the order of 10's of kHz), in order to compare an active spectrum (the system's response to laser driving) with a passive spectrum (the system's response to Brownian motion). In addition, we have an AOD in our detection laser's path, allowing us to calibrate our position detection Quadrant Photo Diode's (QPD) volts to nanometers conversion. Our system allows calibration of every necessary trap parameter in the cell on each individual organelle we trap. We currently trap lipid droplets in Cos-7 cells, and are planning on measuring motor stall forces, motor number, and looking at switching between kinesin based transport and dynein based transport.