Red blood cells are amazingly deformable structures able to recover their initial shape even after large deformations as when passing through tight blood capillaries. The reason for this exceptional property is found in the composition of the membrane and the membrane-cytoskeleton interaction. We investigate the mechanics and the dynamics of RBCs by a unique noninvasive technique, using weak optical tweezers to measure membrane fluctuation amplitudes with μs temporal and sub nm spatial resolution. This enhanced edge detection method allows to span over >4 orders of magnitude in frequency. Hence, we can simultaneously measure red blood cell membrane mechanical properties such as bending modulusκ = 2.8 ± 0.3 × 10−19J = 67.6 ± 7.2 kBT, tension σ = 6.5 ± 2.1 × 10−7N/m, and an effective viscosity ηeff = 81 ± 3.7 × 10−3 Pa s that suggests unknown dissipative processes. We furthermore show that cell mechanics highly depends on the membrane-spectrin interaction mediated by the phosphorylation of the interconnection protein 4.1R. Inhibition and activation of this phosphorylation significantly affects tension and effective viscosity. Our results show that on short time scales (slower than 100 ms) the membrane fluctuates as in thermodynamic equilibrium. At time scales longer than 100 ms, the equilibrium description breaks down and fluctuation amplitudes are higher by 40% than predicted by the membrane equilibrium theory. Possible explanations for this discrepancy are influences of the spectrin that is not included in the membrane theory or nonequilibrium fluctuations that can be accounted for by defining a nonthermal effective energy of up to Eeff = 1.4 ± 0.1 kBT, that corresponds to an actively increased effective temperature.
Wednesday, September 16, 2009
ATP-dependent mechanics of red blood cells
Timo Betz, Martin Lenz, Jean-François Joanny and Cécile Sykes
Tuesday, September 15, 2009
Optical angular momentum transfer to microrotors fabricated by two-photon photopolymerization
Theodor Asavei, Vincent L Y Loke, Marco Barbieri, Timo A Nieminen, Norman R Heckenberg and Halina Rubinsztein-Dunlop
We design, fabricate and test optically driven microrotors a few microns in size. The rotors are trapped and rotated in optical tweezers using an LG02 Laguerre–Gaussian laser beam. We verify that we can accurately measure the total optical torque by measuring the spin angular momentum transfer for three different polarizations, by comparing the optical torque with the optical torque calculated using computational electrodynamics and the viscous drag torque determined from the rotation rate and computational fluid dynamics. The torque agrees with that expected from the design principles and electromagnetic modelling of the torque within the optical trap.
Particle size limits when using optical trapping and deflection of particles for sorting using diode laser bars
Robert W. Applegate Jr., David W. M. Marr, Jeff Squier, and Steven W. Graves
We explore a simple, inexpensive approach to large particle manipulation using diode laser bar optical trapping. This method overcomes limitations that prevent conventional point laser traps from effectively directing large particles. Expanding a previously developed line optical trap model into larger particle regimes, we verify and examine the advantages and limitations of diode laser bar trapping for manipulating particles greater than 100 µm in diameter within fluidic environments for biochemical, biological, and biomedical applications.
We explore a simple, inexpensive approach to large particle manipulation using diode laser bar optical trapping. This method overcomes limitations that prevent conventional point laser traps from effectively directing large particles. Expanding a previously developed line optical trap model into larger particle regimes, we verify and examine the advantages and limitations of diode laser bar trapping for manipulating particles greater than 100 µm in diameter within fluidic environments for biochemical, biological, and biomedical applications.
Thermal motion of a holographically trapped SPM-like probe
Stephen H Simpson and Simon Hanna
By holding a complex object in multiple optical traps, it may be harmonically bound with respect to both its position and its orientation. In this way a small probe, or nanotool, can be manipulated in three dimensions and used to measure and apply directed forces, in the manner of a scanning probe microscope. In this paper we evaluate the thermal motion of such a probe held in holographic optical tweezers, by solving the Langevin equation for the general case of a set of spherical vertices linked by cylindrical rods. The concept of a corner frequency, familiar from the case of an optically trapped sphere, is appropriately extended to represent a set of characteristic frequencies given by the eigenvalues of the product of the stiffness matrix and the inverse hydrodynamic resistance matrix of the tool. These eigenvalues may alternatively be interpreted as inverses of a set of characteristic relaxation times for the system. The approach is illustrated by reference to a hypothetical tool consisting of a triangular arrangement of spheres with a lateral probe. The characteristic frequencies and theoretical resolution of the device are derived; variations of these quantities with tool size and orientation and with the optical power distribution, are also considered.
Monday, September 14, 2009
Combined optical trapping and microphotoluminescence of single InP nanowires
Peter J. Reece, Suriati Paiman, Osama Abdul-Nabi, Qiang Gao, Michael Gal, H. Hoe Tan, and C. Jagadish
In this letter, we demonstrate that microphotoluminescence may be combined with optical trapping for effective optical characterization of single target InP semiconductor nanowires in suspension. Using this technique, we may investigate structural properties of optically trapped nanowires, such as crystalline polytypes and stacking faults. This arrangement may also be used to resolve structural variations along the axis of the trapped nanowire.These results show that photoluminescence measurements may be coupled with optical tweezers without degrading the performance of the optical trap and provide a powerful interrogation tool for preselection of components for nanowire photonic devices.
In this letter, we demonstrate that microphotoluminescence may be combined with optical trapping for effective optical characterization of single target InP semiconductor nanowires in suspension. Using this technique, we may investigate structural properties of optically trapped nanowires, such as crystalline polytypes and stacking faults. This arrangement may also be used to resolve structural variations along the axis of the trapped nanowire.These results show that photoluminescence measurements may be coupled with optical tweezers without degrading the performance of the optical trap and provide a powerful interrogation tool for preselection of components for nanowire photonic devices.
Using optical landscapes to control, direct and isolate aerosol particles
Jon B. Wills, Jason R. Butler, John Palmer and Jonathan P. Reid
We demonstrate the ability to direct the flow of aerosol droplets through a trapping cell using a tailored optical landscape generated by spatial light modulation. Using an optical barrier, droplets held in an optical trap can be effectively isolated from other droplets within the aerosol. To illustrate the effective isolation we compare the influence of different optical landscapes on the flow of free aerosol around a trapped droplet. We also present spectroscopic evidence of the optical barrier effect and apply the technique to permit controlled loading of different aerosol particles into neighbouring optical traps. This method will enable comparative measurements of aerosol properties to be made and facilitate the study of aerosol chemistry in sub-picolitre droplets. It also facilitates the use of an isolated droplet of known composition as a sensitive probe of the gas phase conditions in an aerosol ensemble.
We demonstrate the ability to direct the flow of aerosol droplets through a trapping cell using a tailored optical landscape generated by spatial light modulation. Using an optical barrier, droplets held in an optical trap can be effectively isolated from other droplets within the aerosol. To illustrate the effective isolation we compare the influence of different optical landscapes on the flow of free aerosol around a trapped droplet. We also present spectroscopic evidence of the optical barrier effect and apply the technique to permit controlled loading of different aerosol particles into neighbouring optical traps. This method will enable comparative measurements of aerosol properties to be made and facilitate the study of aerosol chemistry in sub-picolitre droplets. It also facilitates the use of an isolated droplet of known composition as a sensitive probe of the gas phase conditions in an aerosol ensemble.
Optical tweezers with millikelvin precision of temperature-controlled objectives and base-pair resolution
Mohammed Mahamdeh and Erik Schäffer
In optical tweezers, thermal drift is detrimental for highr-esolution measurements. In particular, absorption of the trapping laser light by the microscope objective that focuses the beam leads to heating of the objective and subsequent drift. This entails long equilibration times which may limit sensitive biophysical assays. Here, we introduce an objective temperature feedback system for minimizing thermal drift. We measured that the infrared laser heated the objective by 0.7K per watt of laser power and that the laser focus moved relative to the sample by ≈1 nm/mK due to thermal expansion of the objective. The feedback stabilized the temperature of the trapping objective with millikelvin precision. This enhanced the long-term temperature stability and significantly reduced the settling time of the instrument to about 100 s after a temperature disturbance while preserving single DNA base-pair resolution of surface-coupled assays. Minimizing systematic temperature changes of the objective and concurrent drift is of interest for other high-resolution microscopy techniques. Furthermore, temperature control is often a desirable parameter in biophysical experiments.
In optical tweezers, thermal drift is detrimental for highr-esolution measurements. In particular, absorption of the trapping laser light by the microscope objective that focuses the beam leads to heating of the objective and subsequent drift. This entails long equilibration times which may limit sensitive biophysical assays. Here, we introduce an objective temperature feedback system for minimizing thermal drift. We measured that the infrared laser heated the objective by 0.7K per watt of laser power and that the laser focus moved relative to the sample by ≈1 nm/mK due to thermal expansion of the objective. The feedback stabilized the temperature of the trapping objective with millikelvin precision. This enhanced the long-term temperature stability and significantly reduced the settling time of the instrument to about 100 s after a temperature disturbance while preserving single DNA base-pair resolution of surface-coupled assays. Minimizing systematic temperature changes of the objective and concurrent drift is of interest for other high-resolution microscopy techniques. Furthermore, temperature control is often a desirable parameter in biophysical experiments.
Calculating optical forces using the boundary integral method
Per Jakobsen
In this paper, we show that the boundary integral method is highly efficient for the calculation of optical forces on small dielectric and metallic objects. The boundary integral formulation for the Maxwell equations is stated, and an implementation of the equations is described, tested and used to derive new bistability results for two dielectric spheres in counterpropagating incoherent laser beams.
Thursday, September 10, 2009
Nondimensional analysis of particle behavior during cross-type optical particle separation
Sang Bok Kim, Hyung Jin Sung, and Sang Soo Kim
A nondimensional analysis of particle behavior during cross-type optical particle separation was performed. A new dimensionless number, S, was defined as the ratio of the optical force to the viscous drag force, and the effects of varying S on particle motion were examined. For large S, the particles undergo acceleration, deceleration, and release as they pass through the laser beam. The retention distance is much longer for large S than for small S. In addition, the effects on particle behavior of varying the wavelength of the laser beam, the particle size, and the index of refraction of the particles were investigated. Furthermore, an analytical expression of the retention distance for large S was validated.
A nondimensional analysis of particle behavior during cross-type optical particle separation was performed. A new dimensionless number, S, was defined as the ratio of the optical force to the viscous drag force, and the effects of varying S on particle motion were examined. For large S, the particles undergo acceleration, deceleration, and release as they pass through the laser beam. The retention distance is much longer for large S than for small S. In addition, the effects on particle behavior of varying the wavelength of the laser beam, the particle size, and the index of refraction of the particles were investigated. Furthermore, an analytical expression of the retention distance for large S was validated.
Direct observation of the binding state of the kinesin head to the microtubule
Nicholas R. Guydosh & Steven M. Block
The dimeric motor protein kinesin-1 converts chemical energy from ATP hydrolysis into mechanical work used to transport cargo along microtubules(1,2). Cargo attached to the kinesin stalk moves processively in 8-nm increments(3) as its twin motor domains (heads) carry out an asymmetric, 'hand-over-hand' walk(4-7). The extent of individual head interactions with the microtubule during stepping, however, remains controversial(4,8-14). A major experimental limitation has been the lack of a means to monitor the attachment of an individual head to the microtubule during movement, necessitating indirect approaches. Here we report the development of a single-molecule assay that can directly report head binding in a walking kinesin molecule, and show that only a single head is bound to the microtubule between steps at low ATP concentrations. A bead was linked to one of the two kinesin heads by means of a short DNA tether and used to apply rapidly alternating hindering and assisting loads with an optical trap. The time-dependent difference between forwards and backwards displacements of the bead alternated between two discrete values during stepping, corresponding to those intervals when the linked head adopted a bound or an unbound state. The linked head could only rebind the microtubule once ATP had become bound to its partner head.
The dimeric motor protein kinesin-1 converts chemical energy from ATP hydrolysis into mechanical work used to transport cargo along microtubules(1,2). Cargo attached to the kinesin stalk moves processively in 8-nm increments(3) as its twin motor domains (heads) carry out an asymmetric, 'hand-over-hand' walk(4-7). The extent of individual head interactions with the microtubule during stepping, however, remains controversial(4,8-14). A major experimental limitation has been the lack of a means to monitor the attachment of an individual head to the microtubule during movement, necessitating indirect approaches. Here we report the development of a single-molecule assay that can directly report head binding in a walking kinesin molecule, and show that only a single head is bound to the microtubule between steps at low ATP concentrations. A bead was linked to one of the two kinesin heads by means of a short DNA tether and used to apply rapidly alternating hindering and assisting loads with an optical trap. The time-dependent difference between forwards and backwards displacements of the bead alternated between two discrete values during stepping, corresponding to those intervals when the linked head adopted a bound or an unbound state. The linked head could only rebind the microtubule once ATP had become bound to its partner head.
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