The goal of this project is to develop a new instrument for simultaneous measurements of the structural and the mechanical properties of cytoskeletal protein network in living cells. A working prototype oscillating optical tweezer cytorheometer (OOTC) has already been developed from a work supported by a prior NSF-SGER grant. The present project will be implemented by integrating the existing OOTC with an Olympus spinning disk confocal microscope. The use of optical tweezers to measure mechanical properties of cells has been making significant processes recently. The proposed approach, Oscillating Optical Tweezer Cytorheometer (OOTC), takes advantage of the coherent detection of harmonically modulated particle motions by a lock-in amplifier to increase sensitivity, temporal resolution and simplicity. It has been demonstrated that OOTC can measure the dynamic mechanical modulus in the frequency range of 0.1- 6,000 Hz at a rate as fast as 100 data per second with 1 um3 spatial definition. More importantly, OOTC is capable of distinguishing the intrinsic non-random temporal variations from random fluctuations due to Brownian motion; this capability, not achievable by a conventional approach, is particular useful because living systems are highly dynamic and often exhibit non-thermal, rhythmic behavior. However, as capable as is OOTC, unless we can simultaneously measure the cytoskeletal structures in situ, the mechanical properties data would be as informative as that of "Blind men and the Elephant". Simultaneous and in situ measurements are critical because the polymeric protein network changes shape and reorganizes its structure in time scales from a fraction of a second to hours or days. Advanced optical fluorescent imaging techniques have made it possible to produce 3 dimensional images of the cytoskeletal structure and cytoplasmic components of living cells in great detail by confocal fluorescent microscopy. Integrating OOTC with a confocal microscope will provide eyes to OOTC so we not only feel but also see the cellular structures at the same time. The developed instrument will be used by both undergraduate and graduate students from the physics, biology departments and bioengineering program for education and research. Researchers on and off campus can also use the instrument for studying cell and tissue mechanics problems.
The goal of this project is to develop a new instrument for simultaneous measurements of the structural and the mechanical properties of cytoskeletal protein network in living cells. A working prototype oscillating optical tweezer cytorheometer (OOTC) has already been developed from a work supported by a prior NSF-SGER grant. The present project will be implemented by integrating the existing OOTC with an Olympus spinning disk confocal microscope. Optical tweezers, formed by focused laser beam to hold and manipulate minute organelles in biological cells, have found effective use for measuring intracellular mechanical properties. Using an unique lock-in signal detection scheme, an approach similar to how a car radio pick up music sent by a distant radio station, the proposed approach provides unprecedented sensitivity, temporal resolution and simplicity for measuring the mechanical properties of the cell interior with minimal invasion to living cells. However, as capable as is OOTC, unless one can "see" the cytoskeletal structures in situ, the mechanical properties data would be as informative as that of "Blind men and the Elephant". Advanced fluorescent confocal imaging techniques have made it possible to produce 3 dimensional images of the cytoskeletal structure of living cells in great detail. Integrating OOTC with a confocal microscope will provide eyes to OOTC so we not only feel but also see the cellular structures at the same time. The developed instrument will be used by both undergraduate and graduate students from the physics, biology departments and bioengineering program for education and research. Researchers on and off campus can also use the instrument for studying cell and tissue mechanics problems.
