We present a system employing a dynamic diffractive optical element to control properties of two counterpropagating beams overlapping within a sample chamber. This system allows us to eliminate optical aberrations along both beam pathways and arbitrarily switch between various numbers of laser beams and their spatial profiles (i.e. Gaussian, Laguerre-Gaussian, Bessel beams, etc.). We successfully tested various counter-propagating dual-beam configurations including optical manipulation of both high and low index particles in water or air, particle delivery in an optical conveyor belt and the formation of colloidal solitons by optical binding. Furthermore, we realized a novel optical mixer created by particles spiraling in counter-propagating interfering optical vortices and a new tool for optical tomography or localized spectroscopy enabling sterile contactless rotation and reorientation of a trapped living cell.
Tuesday, March 15, 2011
The holographic optical micro-manipulation system based on counter-propagating beams
T. Čižmár, O. Brzobohatý, K. Dholakia, P. Zemánek
We present a system employing a dynamic diffractive optical element to control properties of two counterpropagating beams overlapping within a sample chamber. This system allows us to eliminate optical aberrations along both beam pathways and arbitrarily switch between various numbers of laser beams and their spatial profiles (i.e. Gaussian, Laguerre-Gaussian, Bessel beams, etc.). We successfully tested various counter-propagating dual-beam configurations including optical manipulation of both high and low index particles in water or air, particle delivery in an optical conveyor belt and the formation of colloidal solitons by optical binding. Furthermore, we realized a novel optical mixer created by particles spiraling in counter-propagating interfering optical vortices and a new tool for optical tomography or localized spectroscopy enabling sterile contactless rotation and reorientation of a trapped living cell.
We present a system employing a dynamic diffractive optical element to control properties of two counterpropagating beams overlapping within a sample chamber. This system allows us to eliminate optical aberrations along both beam pathways and arbitrarily switch between various numbers of laser beams and their spatial profiles (i.e. Gaussian, Laguerre-Gaussian, Bessel beams, etc.). We successfully tested various counter-propagating dual-beam configurations including optical manipulation of both high and low index particles in water or air, particle delivery in an optical conveyor belt and the formation of colloidal solitons by optical binding. Furthermore, we realized a novel optical mixer created by particles spiraling in counter-propagating interfering optical vortices and a new tool for optical tomography or localized spectroscopy enabling sterile contactless rotation and reorientation of a trapped living cell.
Monday, March 14, 2011
Brownian motion in a Maxwell fluid
Matthias Grimm, Sylvia Jeney and Thomas Franosch
The equilibrium dynamics of a spherical particle immersed in a complex Maxwell fluid is analyzed in terms of velocity autocorrelation function (VACF), mean-square displacement (MSD), and power spectral density (PSD). We elucidate the role of hydrodynamic memory and its interplay with medium viscoelasticity for a free and a harmonically confined particle. The elastic response at high frequencies introduces oscillations in the VACF, which are found to be strongly damped by the coupling to the fluid. We show that in all Maxwell fluids hydrodynamic memory eventually leads to a power-law decay in the VACF as is already known for Newtonian fluids. The MSD displays asymptotically an intermediate plateau reflecting the elastic restoring forces of the medium. In the frequency domain, the PSD exhibits at high frequencies a step due to the trapping, whereas the low-frequency decay reflects the viscoelastic relaxation. Our results suggest that high-frequency microrheology is well-suited to infer the elastic modulus, which is sensitive over a wide range of Maxwell times.
DOI
The equilibrium dynamics of a spherical particle immersed in a complex Maxwell fluid is analyzed in terms of velocity autocorrelation function (VACF), mean-square displacement (MSD), and power spectral density (PSD). We elucidate the role of hydrodynamic memory and its interplay with medium viscoelasticity for a free and a harmonically confined particle. The elastic response at high frequencies introduces oscillations in the VACF, which are found to be strongly damped by the coupling to the fluid. We show that in all Maxwell fluids hydrodynamic memory eventually leads to a power-law decay in the VACF as is already known for Newtonian fluids. The MSD displays asymptotically an intermediate plateau reflecting the elastic restoring forces of the medium. In the frequency domain, the PSD exhibits at high frequencies a step due to the trapping, whereas the low-frequency decay reflects the viscoelastic relaxation. Our results suggest that high-frequency microrheology is well-suited to infer the elastic modulus, which is sensitive over a wide range of Maxwell times.
DOI
Red blood cell dynamics: from spontaneous fluctuations to non-linear response
Young Zoon Yoon, Jurij Kotar, Aidan T. Brown and Pietro Cicuta
We studied experimentally the mechanical properties of the red blood cell. By attaching beads biochemically on the cell membrane at diametrically opposite positions, the membrane movements can be detected very accurately, and a deformation of the cell can be imposed. A measurement of the mechanical properties at very small amplitudes is obtained by fluctuation analysis, and compared to the stiffness at larger deformations, obtained by stretching the cells via optical traps whilst monitoring the force. The cells are also probed at various conditions of pre-strain. These measurements show clearly a stiffening with strain and with pre-strain, which is strongest at low frequencies of deformation. The cell is measured to be slightly softer from fluctuation analysis, but consistent simply with the fact that the oscillation amplitude in fluctuations is very small. There is no evidence in these experiments of non-thermal sources of membrane motion, although non-thermal noise may be present within experimental error.
DOI
We studied experimentally the mechanical properties of the red blood cell. By attaching beads biochemically on the cell membrane at diametrically opposite positions, the membrane movements can be detected very accurately, and a deformation of the cell can be imposed. A measurement of the mechanical properties at very small amplitudes is obtained by fluctuation analysis, and compared to the stiffness at larger deformations, obtained by stretching the cells via optical traps whilst monitoring the force. The cells are also probed at various conditions of pre-strain. These measurements show clearly a stiffening with strain and with pre-strain, which is strongest at low frequencies of deformation. The cell is measured to be slightly softer from fluctuation analysis, but consistent simply with the fact that the oscillation amplitude in fluctuations is very small. There is no evidence in these experiments of non-thermal sources of membrane motion, although non-thermal noise may be present within experimental error.
DOI
Downstream DNA Tension Regulates the Stability of the T7 RNA Polymerase Initiation Complex
Gary M. Skinner, Bennett S. Kalafut and Koen Visscher
Gene transcription by the enzyme RNA polymerase is tightly regulated. In many cases, such as in the lac operon in Escherichia coli, this regulation is achieved through the action of protein factors on DNA. Because DNA is an elastic polymer, its response to enzymatic processing can lead to mechanical perturbations (e.g., linear stretching and supercoiling) that can affect the operation of other DNA processing complexes acting elsewhere on the same substrate molecule. Using an optical-tweezers assay, we measured the binding kinetics between single molecules of bacteriophage T7 RNA polymerase and DNA, as a function of tension. We found that increasing DNA tension under conditions that favor formation of the open complex results in destabilization of the preinitiation complex. Furthermore, with zero ribonucleotides present, when the closed complex is favored, we find reduced tension sensitivity, implying that it is predominantly the open complex that is sensitive. This result strongly supports the “scrunching” model for T7 transcription initiation, as the applied tension acts against the movement of the DNA into the scrunched state, and introduces linear DNA tension as a potential regulatory quantity for transcription initiation.
DOI
Gene transcription by the enzyme RNA polymerase is tightly regulated. In many cases, such as in the lac operon in Escherichia coli, this regulation is achieved through the action of protein factors on DNA. Because DNA is an elastic polymer, its response to enzymatic processing can lead to mechanical perturbations (e.g., linear stretching and supercoiling) that can affect the operation of other DNA processing complexes acting elsewhere on the same substrate molecule. Using an optical-tweezers assay, we measured the binding kinetics between single molecules of bacteriophage T7 RNA polymerase and DNA, as a function of tension. We found that increasing DNA tension under conditions that favor formation of the open complex results in destabilization of the preinitiation complex. Furthermore, with zero ribonucleotides present, when the closed complex is favored, we find reduced tension sensitivity, implying that it is predominantly the open complex that is sensitive. This result strongly supports the “scrunching” model for T7 transcription initiation, as the applied tension acts against the movement of the DNA into the scrunched state, and introduces linear DNA tension as a potential regulatory quantity for transcription initiation.
DOI
Dissociation of Bimolecular αIIbβ3-Fibrinogen Complex under a Constant Tensile Force
Rustem I. Litvinov, Valeri Barsegov, Andrew J. Schissler, Andrew R. Fisher, Joel S. Bennett, John W. Weisel and Henry Shuman
The regulated ability of integrin αIIbβ3 to bind fibrinogen plays a crucial role in platelet aggregation, adhesion, and hemostasis. Employing an optical-trap-based electronic force clamp, we studied the thermodynamics and kinetics of αIIbβ3-fibrinogen bond formation and dissociation under constant unbinding forces, mimicking the forces of physiologic blood shear on a thrombus. The distribution of bond lifetimes was bimodal, indicating that the αIIbβ3-fibrinogen complex exists in two bound states with different mechanical stability. The αIIbβ3 antagonist, abciximab, inhibited binding without affecting the unbinding kinetics, whereas Mn2+ biased the αIIbβ3-fibrinogen complex to the strong bound state with reduced off-rate. The average bond lifetimes decreased exponentially with increasing pulling force from
5 pN to 50 pN, suggesting that in this force range the αIIbβ3-fibrinogen interactions are classical slip bonds. We found no evidence for catch bonds, which is consistent with the known lack of shear-enhanced platelet adhesion on fibrinogen-coated surfaces. Taken together, these data provide important quantitative and qualitative characteristics of αIIbβ3-fibrinogen binding and unbinding that underlie the dynamics of platelet adhesion and aggregation in blood flow.
DOI
The regulated ability of integrin αIIbβ3 to bind fibrinogen plays a crucial role in platelet aggregation, adhesion, and hemostasis. Employing an optical-trap-based electronic force clamp, we studied the thermodynamics and kinetics of αIIbβ3-fibrinogen bond formation and dissociation under constant unbinding forces, mimicking the forces of physiologic blood shear on a thrombus. The distribution of bond lifetimes was bimodal, indicating that the αIIbβ3-fibrinogen complex exists in two bound states with different mechanical stability. The αIIbβ3 antagonist, abciximab, inhibited binding without affecting the unbinding kinetics, whereas Mn2+ biased the αIIbβ3-fibrinogen complex to the strong bound state with reduced off-rate. The average bond lifetimes decreased exponentially with increasing pulling force from
5 pN to 50 pN, suggesting that in this force range the αIIbβ3-fibrinogen interactions are classical slip bonds. We found no evidence for catch bonds, which is consistent with the known lack of shear-enhanced platelet adhesion on fibrinogen-coated surfaces. Taken together, these data provide important quantitative and qualitative characteristics of αIIbβ3-fibrinogen binding and unbinding that underlie the dynamics of platelet adhesion and aggregation in blood flow.DOI
Optically Directed Assembly of Continuous Mesoscale Filaments
J. T. Bahns, S. K. R. S. Sankaranarayanan, S. K. Gray, and L. Chen
We demonstrate irreversible continuous filament formation when a weak laser focus is positioned near the edge of an evaporating colloidal droplet containing carbon and gold nanoparticles. Optical trapping, hydrothermal, and chemical interactions lead to controlled colloidal synthesis of stable, irreversible mesoscale filaments of arbitrary shape and size. Mechanisms for this optically directed assembly are discussed with fluid dynamics, molecular dynamics, and lattice kinetic Monte Carlo calculations.
DOI
We demonstrate irreversible continuous filament formation when a weak laser focus is positioned near the edge of an evaporating colloidal droplet containing carbon and gold nanoparticles. Optical trapping, hydrothermal, and chemical interactions lead to controlled colloidal synthesis of stable, irreversible mesoscale filaments of arbitrary shape and size. Mechanisms for this optically directed assembly are discussed with fluid dynamics, molecular dynamics, and lattice kinetic Monte Carlo calculations.
DOI
Sunday, March 13, 2011
Optical tweezers and paradoxes in electromagnetism
Robert N C Pfeifer, Timo A Nieminen, Norman R Heckenberg and Halina Rubinsztein-Dunlop
The widespread application of optical forces and torques has contributed to renewed interest in the fundamentals of the electromagnetic force and torque, including long-standing paradoxes such as the Abraham–Minkowski controversy and the angular momentum density of a circularly polarized plane wave. We discuss the relationship between these electromagnetic paradoxes and optical tweezers. In particular, consideration of possible optical tweezers experiments to attempt to resolve these paradoxes strongly suggests that they are beyond experimental resolution, yielding identical observable results in all cases.
DOI
The widespread application of optical forces and torques has contributed to renewed interest in the fundamentals of the electromagnetic force and torque, including long-standing paradoxes such as the Abraham–Minkowski controversy and the angular momentum density of a circularly polarized plane wave. We discuss the relationship between these electromagnetic paradoxes and optical tweezers. In particular, consideration of possible optical tweezers experiments to attempt to resolve these paradoxes strongly suggests that they are beyond experimental resolution, yielding identical observable results in all cases.
DOI
Stereoscopic particle tracking for 3D touch, vision and closed-loop control in optical tweezers
Richard Bowman, Daryl Preece, Graham Gibson and Miles Padgett
Force measurement in an interactive 3D micromanipulation system can allow the user to make delicate adjustments, and to explore surfaces with touch as well as vision. We present a system to achieve this on the micron scale using stereoscopic particle tracking combined with holographic optical tweezers, which can track particles with nanometre accuracy. 2D tracking of particles in each of the stereo images gives 3D positions for each particle. This takes less than 200 µs per image pair, using a 1D 'symmetry transform' applied to each row and column of a 2D image, which can maintain tracking of particles throughout the 10 µm axial range. The only parameters required are the geometry of the imaging system, and therefore there is no need to recalibrate for different particle sizes or refractive indices. Consequently, we can calculate the force exerted by the optical trap in real time at 1 kilohertz, allowing us to implement a force-feedback interface (with a loop rate of 400 Hz). In combination with our OpenGL hologram calculation engine, the system has a closed-loop bandwidth of 20 Hz. This allows us to stabilize trapped particles axially through active feedback, cancelling out some Brownian motion. For the weak traps we use here (spring constant k≈2 pN µm − 1), this results in a threefold increase in axial stiffness. We demonstrate the 3D interface by probing an oil droplet, mapping out its surface in the y–z plane.
DOI
Force measurement in an interactive 3D micromanipulation system can allow the user to make delicate adjustments, and to explore surfaces with touch as well as vision. We present a system to achieve this on the micron scale using stereoscopic particle tracking combined with holographic optical tweezers, which can track particles with nanometre accuracy. 2D tracking of particles in each of the stereo images gives 3D positions for each particle. This takes less than 200 µs per image pair, using a 1D 'symmetry transform' applied to each row and column of a 2D image, which can maintain tracking of particles throughout the 10 µm axial range. The only parameters required are the geometry of the imaging system, and therefore there is no need to recalibrate for different particle sizes or refractive indices. Consequently, we can calculate the force exerted by the optical trap in real time at 1 kilohertz, allowing us to implement a force-feedback interface (with a loop rate of 400 Hz). In combination with our OpenGL hologram calculation engine, the system has a closed-loop bandwidth of 20 Hz. This allows us to stabilize trapped particles axially through active feedback, cancelling out some Brownian motion. For the weak traps we use here (spring constant k≈2 pN µm − 1), this results in a threefold increase in axial stiffness. We demonstrate the 3D interface by probing an oil droplet, mapping out its surface in the y–z plane.
DOI
Optimizing active and passive calibration of optical tweezers
M Andersson, F Czerwinski and L B Oddershede
To obtain quantitative information from optical trapping experiments it is essential to perform a precise force calibration. Therefore, sources of noise should be pinpointed and eliminated. Fourier analysis is routinely used to calibrate optical trapping assays because it is excellent for pinpointing high frequency noise. In addition, Allan variance analysis is particularly useful for quantifying low frequency noise and for predicting the optimal measurement time. We show how to use Allan variance in combination with Fourier analysis for optimal calibration and noise reduction in optical trapping assays. The methods are applied to passive assays, utilizing the thermal motion of a trapped particle, and to active assays where the bead is harmonically driven. The active method must be applied in assays where, for example, the viscoelastic properties of the medium or the size or shape of the trapped object are unknown. For measurement times shorter than the optimal calibration time the noise is larger in active than in the passive assays. For times equal to or longer than the optimal measurement time, though, the noise on passive and active assays is identical. As an example, we show how to quantify the influence on measurement noise of bead size and chamber geometry in active and passive assays.
DOI
To obtain quantitative information from optical trapping experiments it is essential to perform a precise force calibration. Therefore, sources of noise should be pinpointed and eliminated. Fourier analysis is routinely used to calibrate optical trapping assays because it is excellent for pinpointing high frequency noise. In addition, Allan variance analysis is particularly useful for quantifying low frequency noise and for predicting the optimal measurement time. We show how to use Allan variance in combination with Fourier analysis for optimal calibration and noise reduction in optical trapping assays. The methods are applied to passive assays, utilizing the thermal motion of a trapped particle, and to active assays where the bead is harmonically driven. The active method must be applied in assays where, for example, the viscoelastic properties of the medium or the size or shape of the trapped object are unknown. For measurement times shorter than the optimal calibration time the noise is larger in active than in the passive assays. For times equal to or longer than the optimal measurement time, though, the noise on passive and active assays is identical. As an example, we show how to quantify the influence on measurement noise of bead size and chamber geometry in active and passive assays.
DOI
Agglutination of Histoplasma capsulatum by IgG Monoclonal Antibodies against Hsp60 Impacts Macrophage Effector Functions
Allan Jefferson Guimarães, Susana Frases, Bruno Pontes, Mariana Duarte de Cerqueira, Marcio L. Rodrigues, Nathan Bessa Viana, Leonardo Nimrichter, and Joshua Daniel Nosanchuk
Histoplasma capsulatum can efficiently survive within macrophages, facilitating H. capsulatum translocation from the lung into the lymphatics and bloodstream. We have recently generated monoclonal antibodies (MAbs) to an H. capsulatum surface-expressed heat shock protein of 60 kDa (Hsp60) that modify disease in a murine histoplasmosis model. Interestingly, the MAbs induced different degrees of yeast cell agglutination in vitro. In the present study, we characterized the agglutination effects of the antibodies to Hsp60 on H. capsulatum yeast cells by light microscopy, flow cytometry, dynamic light scattering, measuring zeta potential, and using optical tweezers. We found that immunoglobulin Gs (IgGs) to Hsp60 cause H. capsulatum aggregation dependent on the (i) concentration of MAbs, (ii) MAb binding constant, and (iii) IgG subclass. Furthermore, infection of macrophages using agglutinates of various sizes after incubation with different Hsp60-binding MAbs induced association to macrophages through distinct cellular receptors and differentially affected macrophage antifungal functions. Hence, the capacity of IgG MAbs to agglutinateH. capsulatum significantly impacted pathogenic mechanisms of H. capsulatum during macrophage infection, and the effect was dependent on the antibody subclass and antigen epitope.
DOI
Histoplasma capsulatum can efficiently survive within macrophages, facilitating H. capsulatum translocation from the lung into the lymphatics and bloodstream. We have recently generated monoclonal antibodies (MAbs) to an H. capsulatum surface-expressed heat shock protein of 60 kDa (Hsp60) that modify disease in a murine histoplasmosis model. Interestingly, the MAbs induced different degrees of yeast cell agglutination in vitro. In the present study, we characterized the agglutination effects of the antibodies to Hsp60 on H. capsulatum yeast cells by light microscopy, flow cytometry, dynamic light scattering, measuring zeta potential, and using optical tweezers. We found that immunoglobulin Gs (IgGs) to Hsp60 cause H. capsulatum aggregation dependent on the (i) concentration of MAbs, (ii) MAb binding constant, and (iii) IgG subclass. Furthermore, infection of macrophages using agglutinates of various sizes after incubation with different Hsp60-binding MAbs induced association to macrophages through distinct cellular receptors and differentially affected macrophage antifungal functions. Hence, the capacity of IgG MAbs to agglutinateH. capsulatum significantly impacted pathogenic mechanisms of H. capsulatum during macrophage infection, and the effect was dependent on the antibody subclass and antigen epitope.
DOI
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