Optical tweezers have proven themselves as an indispensable tool in modern biology. By using momentum imparted by laser beams onto micrometer sized beads, optical tweezers can precisely position and apply forces on the trapped objects. The technique is immensely useful for the study of biological molecules, living cells, and other micron-sized objects as distances in the order of nanometer scale and small forces at pico-newtons can be measured with high precision, which are otherwise difficult to measure.
One way to use optical tweezers is to apply stretching and translational forces on chain-like biopolymers such as enzymes, RNA, and DNA molecules. By accurately measuring the forces and positions required to manipulate the molecule into its different conformational states, important conclusions can be made about the chemistry, folding parameters, geometry, and energy barriers that are vital for the biological function. The stretching is achieved by anchoring both ends of the molecule to micron sized polystyrene beads. One of these polystyrene beads is held in place at the end of a micropipette while the other is suspended and trapped with the tweezers instrument.
One impediment to such discoveries is the low rate at which data is collected. Sparse data collection rate makes analysis of high frequency fast kinetics data difficult. Many of the existing data sets generated from optical tweezers contain these high frequency features like sharp peaks and rapid transitions to intermediate positions. By increasing the data collection rate details in these peaks and sharp transitions can be addressed. This project seeks to increase the existing data collection rate from 4 kHz to 20 kHz. This will lead to a clearer understanding of the molecular processes at hand and open up new areas of investigation.
Currently, the tweezers instrument in the Mandal Lab uses series of electronic boards for analog/digital signal conversion, force and position measurements from various position sensing devices (PSDs), piezo drivers, feedback control of laser traps, and motor controllers. While electronic boards provide a better resolution of bead position over many of the other methods such as video microscopy, the limit is set by the virtual serial port over USB. The main components that will make this high speed data acquisition possible is by replacing the electronic boards with a National Instruments (NI) Data Acquisition (DAQ) System. LabView software will provide a user friendly interface that runs on a vibration free desktop computer. This setup must also be thoroughly documented, easy to use, and readily expandable in the future. LabView software and NI DAQ hardware is designed and industry-proven for high-speed data acquisition.
The project will begin by interfacing to NI hardware to the existing DAQ system to access the signals to be logged. This will allow both systems to run simultaneously in order to verify the accuracy of the new NI DAQ system. Initially, passive recording of signals corresponding to the force and position of trapped objected will be attempted. Next, the DAQ system will use its analog-voltage-output capability to control the optics that positions the trapped object. This phase also requires an investigation of laser beam steering mechanisms like piezo-electrically/acousto-optically actuated mirrors. The selected system must be fast enough for quickly recurring movement of the trapped object. The final stage would be using the digital I/O capability of the NI DAQ system to control the motorized three-axis translational movement of the sample chamber. This movement allows for rough positioning of the sample while manipulating the laser beams that trap the polystyrene bead allows for fine nanometer-scale control.
Optical tweezers are essential for studying and manipulating the microscopic objects frequently encountered in biological systems. By increasing the rate of data collection of this instrument , this project hopes to set the stage for future advances in this field.