With suitable calibration, optical tweezers can be used to measure forces. If the maximum force that can be exerted is of interest, calibration can be performed using viscous drag to remove a particle from the trap, typically by moving the stage. The stage velocity required to remove the particle then gives the escape force. However, the escape force can vary by up to 30% or more, depending on the particle trajectory. This can have significant quantitative impact on measurements. We describe the variation of escape force and escape trajectory, using both experimental measurements and simulations, and discuss implications for experimental measurement of forces.
We have estimated the mitotic forces exerted on individual isolated mammalian chromosomes using optical trapping. The chromosomes were trapped by an optical tweezers system created by a continuous wave ytterbium laser at 1064 nm. Individual chromosomes were trapped at different in situ powers in the range of ≈20-50 mW. The corresponding trapping forces were determined by a viscous drag method. In the range of laser powers used, the preliminary data show a linear relationship between the chromosome trapping forces and in situ powers. We have calculated the dimensionless trapping efficiency coefficient (Q) of the chromosomes at 1064 nm and the corresponding effects of trapping power on Q. The value of Q in our experiments was determined to be ≈0.01. The results of this study validate optical tweezers as a non-invasive and precise technique to determine intracellular forces in general, and specifically, the spindle forces exerted on the chromosomes during cell division.
In this study, we investigated the effects of size and surrounding media viscosity on trapping of microspheres. A continuous wave ytterbium fiber laser with a 1064 nm wavelength was used to create an optical tweezers system for optical manipulation experiments. Briefly, the system consisted of an inverted microscope, and a 100X 1.4 NA oil immersion objective through which the laser beam converged to form the optical trap. The laser beam was collimated, steered, and coupled to the microscope through the epifluorescence microscope port. The laser power at the trap focal spot was determined by measuring the input power at the back aperture of the objective multiplied by the objective transmission factor at 1064 nm measured by a modified dual objective method. Polystyrene microspheres varying in diameter from 5 to 15 microns were suspended in liquid media in glass bottom petri dishes prior to trapping experiments. The microspheres were trapped at different trapping powers, and fluidic viscous drag forces where applied to the optically trapped microspheres by driving a computer controlled 2D motorized microscope stage at known velocities. The drag forces were calculated at the point that the microspheres fell out of the trap, based on the Stokes equation for flow around spheres. The data show a linear relationship between trapping force and trap power within the range of the microsphere diameters and media viscosity values used. The work includes calculation of the dimensionless trap efficiency coefficient (Q) at 1064 nm wavelength and the corresponding effects of media viscosity and microsphere size on (Q).
In this study, we investigated nanomechanical properties of cell membranes in response to elongation at different rates. Membrane nanotubes (tethers) were pulled at different pulling rates by an optically-trapped fluorescent microsphere as recorded and analyzed for low (1 μm/s) and high (100 μm/s) pulling rates. The force relaxation response of membrane nanotubes exhibited a bi-phasic behavior including fast and slow relaxation processes at low and high pulling rates. The fast and slow force relaxation time constants were 0.388±0.21 s and 11.74±3.35 s, in response to pulling rate of 1 μm/s, respectively and significantly decreased at higher pulling rates. These reductions in the time constants are suggestive of reduced viscous effects and weakened adhesions between the membrane and the cytoskeleton during rapid pulling.