Computer-automated program for calibration of optical tweezers

Abstract

An optical tweezers (OT) system uses focused laser light to contain and manipulate nano-scale to micro-scale particles. Trap stiffness is the quantitative measurement of the ability to trap a particle. For some techniques, this measurement depends on an accurate knowledge of the particle's position in time. A position sensing detector (PSD) is used to track particle motion by detecting laser light from the trapping region. The PSD outputs voltages corresponding to the x- and y-coordinates of particle motion, providing a means of knowing the location of the particle in time. An OT system requires a calibration to convert the measured voltages into accurate distances. This process is time-consuming and frequently needs to be repeated, however, with the growing availability of computer-aided data acquisition and control, the complete process can now be automated, reducing time spent by researchers and increasing level of accuracy of future measurements. We have developed a program written in LabVIEW that will, after initialization, 1) via image processing, calibrate the pixel size of the camera, 2) calibrate the optical tweezer position detector by controlling a motorized mirror to move a trapped bead through a detection laser with simultaneous position detector signal measurements, 3) re-align the trap beam and the detection beam by motorized mirror control, 4) measure position data for the same trapped particle being illuminated by the detection beam, and 5) analyze the position signal via the power spectrum method and equipartition method to give two trap stiffness values for comparison. Previous automated calibration methods require additional and sometimes costly equipment as well as some precalibration of stage motion or pixel size. Here, the user only needs to input the known size of the bead (provided by the manufacturer) into the program, insert their prepared slide into their microscope, input some parameters and make selections, and click "start" in order to achieve experimental values of the camera and position detector calibrations, as well as trap stiffness. We intend to implement many other calibration techniques that require additional equipment, but have designed this initial system for use in a standard position-detection-capable OT setup as long as it has a digital camera and motorized mirror that can be controlled with LabVIEW.