Karen M. Ridge, Dale Shumaker, Amélie Robert, Caroline Hookway, Vladimir I. Gelfand, Paul A. Janmey, Jason Lowery, Ming Guo, David A. Weitz, Edward Kuczmarski, Robert D. Goldman
The type III intermediate filament protein vimentin was once thought to function mainly as a static structural protein in the cytoskeleton of cells of mesenchymal origin. Now, however, vimentin is known to form a dynamic, flexible network that plays an important role in a number of signaling pathways. Here, we describe various methods that have been developed to investigate the cellular functions of the vimentin protein and intermediate filament network, including chemical disruption, photoactivation and photoconversion, biolayer interferometry, soluble bead binding assay, three-dimensional substrate experiments, collagen gel contraction, optical-tweezer active microrheology, and force spectrum microscopy. Using these techniques, the contributions of vimentin to essential cellular processes can be probed in ever further detail.
Wednesday, December 30, 2015
Optical trapping performance of dielectric-metallic patchy particles
Joseph L. Lawson, Nathan J. Jenness, and Robert L. Clark
We demonstrate a series of simulation experiments examining the optical trapping behavior of composite micro-particles consisting of a small metallic patch on a spherical dielectric bead. A full parameter space of patch shapes, based on current state of the art manufacturing techniques, and optical properties of the metallic film stack is examined. Stable trapping locations and optical trap stiffness of these particles are determined based on the particle design and potential particle design optimizations are discussed. A final test is performed examining the ability to incorporate these composite particles with standard optical trap metrology technologies.
DOI
We demonstrate a series of simulation experiments examining the optical trapping behavior of composite micro-particles consisting of a small metallic patch on a spherical dielectric bead. A full parameter space of patch shapes, based on current state of the art manufacturing techniques, and optical properties of the metallic film stack is examined. Stable trapping locations and optical trap stiffness of these particles are determined based on the particle design and potential particle design optimizations are discussed. A final test is performed examining the ability to incorporate these composite particles with standard optical trap metrology technologies.
DOI
Tuesday, December 29, 2015
Charged hydrophobic colloids at an oil–aqueous phase interface
Colm P. Kelleher, Anna Wang, Guillermo Iván Guerrero-García, Andrew D. Hollingsworth, Rodrigo E. Guerra, Bhaskar Jyoti Krishnatreya, David G. Grier, Vinothan N. Manoharan, and Paul M. Chaikin
Hydrophobic poly(methyl methacrylate) (PMMA) colloidal particles, when dispersed in oil with a relatively high dielectric constant, can become highly charged. In the presence of an interface with a conducting aqueous phase, image-charge effects lead to strong binding of colloidal particles to the interface, even though the particles are wetted very little by the aqueous phase. We study both the behavior of individual colloidal particles as they approach the interface and the interactions between particles that are already interfacially bound. We demonstrate that using particles which are minimally wetted by the aqueous phase allows us to isolate and study those interactions which are due solely to charging of the particle surface in oil. Finally, we show that these interactions can be understood by a simple image-charge model in which the particle charge q is the sole fitting parameter.
DOI
Hydrophobic poly(methyl methacrylate) (PMMA) colloidal particles, when dispersed in oil with a relatively high dielectric constant, can become highly charged. In the presence of an interface with a conducting aqueous phase, image-charge effects lead to strong binding of colloidal particles to the interface, even though the particles are wetted very little by the aqueous phase. We study both the behavior of individual colloidal particles as they approach the interface and the interactions between particles that are already interfacially bound. We demonstrate that using particles which are minimally wetted by the aqueous phase allows us to isolate and study those interactions which are due solely to charging of the particle surface in oil. Finally, we show that these interactions can be understood by a simple image-charge model in which the particle charge q is the sole fitting parameter.
DOI
Use of Raman optical tweezers for cell cycle analysis
Sunita Ahlawat, Aniket Chowdhury, Abha Uppal, Nitin Kumar and Pradeep Kumar Gupta
We report the results of our investigations on the use of Raman optical tweezers for label free analysis of cells in different phases of their cell cycle. The studies performed on human colon adenocarcinoma (Colo-205) cells synchronized in G0/G1 and G2/M phases showed that the DNA Raman band at 783 cm-1 in the Raman spectra of optically trapped cells can provide information about the DNA content in the nucleus of the cell without the need for isolation of nucleus. The histograms of intensity of this band among the cell populations were found to corroborate the results obtained from fluorescence image cytometry performed on DAPI stained cells.
DOI
We report the results of our investigations on the use of Raman optical tweezers for label free analysis of cells in different phases of their cell cycle. The studies performed on human colon adenocarcinoma (Colo-205) cells synchronized in G0/G1 and G2/M phases showed that the DNA Raman band at 783 cm-1 in the Raman spectra of optically trapped cells can provide information about the DNA content in the nucleus of the cell without the need for isolation of nucleus. The histograms of intensity of this band among the cell populations were found to corroborate the results obtained from fluorescence image cytometry performed on DAPI stained cells.
DOI
Spinning of a submicron sphere by Airy beams
Kyoung-Youm Kim and Saehwa Kim
We show that by employing two incoherent counter-propagating Airy beams, we can manipulate a submicron sphere to spin around a transverse axis. We can control not only the spinning speed, but also the direction of the spinning axis by changing the polarization directions of Airy beams.
DOI
We show that by employing two incoherent counter-propagating Airy beams, we can manipulate a submicron sphere to spin around a transverse axis. We can control not only the spinning speed, but also the direction of the spinning axis by changing the polarization directions of Airy beams.
DOI
Contact efflorescence as a pathway for crystallization of atmospherically relevant particles
Ryan D. Davis, Sara Lance, Joshua A. Gordon, Shuichi B. Ushijima, and Margaret A. Tolbert
Inadequate knowledge of the phase state of atmospheric particles represents a source of uncertainty in global climate and air quality models. Hygroscopic aqueous inorganic particles are often assumed to remain liquid throughout their atmospheric lifetime or only (re)crystallize at low relative humidity (RH) due to the kinetic limitations of efflorescence (salt crystal nucleation and growth from an aqueous solution). Here we present experimental observations of a previously unexplored heterogeneous nucleation pathway that we have termed “contact efflorescence,” which describes efflorescence initiated by an externally located solid particle coming into contact with the surface of a metastable aqueous microdroplet. This study demonstrates that upon a single collision, contact efflorescence is a pathway for crystallization of atmospherically relevant aqueous particles at high ambient RH (≤80%). Soluble inorganic crystalline particles were used as contact nuclei to induce efflorescence of aqueous ammonium sulfate [(NH4)2SO4], sodium chloride (NaCl), and ammonium nitrate (NH4NO3), with efflorescence being observed in several cases close to their deliquescence RH values (80%, 75%, and 62%, respectively). To our knowledge, these observations represent the highest reported efflorescence RH values for microdroplets of these salts. These results are particularly important for considering the phase state of NH4NO3, where the contact efflorescence RH (∼20–60%) is in stark contrast to the observation that NH4NO3 microdroplets do not homogeneously effloresce, even when exposed to extremely arid conditions (<1% RH). Considering the occurrence of particle collisions in the atmosphere (i.e., coagulation), these observations of contact efflorescence challenge many assumptions made about the phase state of inorganic aerosol.
DOI
Inadequate knowledge of the phase state of atmospheric particles represents a source of uncertainty in global climate and air quality models. Hygroscopic aqueous inorganic particles are often assumed to remain liquid throughout their atmospheric lifetime or only (re)crystallize at low relative humidity (RH) due to the kinetic limitations of efflorescence (salt crystal nucleation and growth from an aqueous solution). Here we present experimental observations of a previously unexplored heterogeneous nucleation pathway that we have termed “contact efflorescence,” which describes efflorescence initiated by an externally located solid particle coming into contact with the surface of a metastable aqueous microdroplet. This study demonstrates that upon a single collision, contact efflorescence is a pathway for crystallization of atmospherically relevant aqueous particles at high ambient RH (≤80%). Soluble inorganic crystalline particles were used as contact nuclei to induce efflorescence of aqueous ammonium sulfate [(NH4)2SO4], sodium chloride (NaCl), and ammonium nitrate (NH4NO3), with efflorescence being observed in several cases close to their deliquescence RH values (80%, 75%, and 62%, respectively). To our knowledge, these observations represent the highest reported efflorescence RH values for microdroplets of these salts. These results are particularly important for considering the phase state of NH4NO3, where the contact efflorescence RH (∼20–60%) is in stark contrast to the observation that NH4NO3 microdroplets do not homogeneously effloresce, even when exposed to extremely arid conditions (<1% RH). Considering the occurrence of particle collisions in the atmosphere (i.e., coagulation), these observations of contact efflorescence challenge many assumptions made about the phase state of inorganic aerosol.
DOI
Monday, December 28, 2015
Development and characterization of a eukaryotic expression system for human type II procollagen
Andrew Wieczorek, Naghmeh Rezaei, Clara K. Chan, Chuan Xu, Preety Panwar, Dieter Brömme, Erika F. Merschrod S. and Nancy R. Forde
Triple helical collagens are the most abundant structural protein in vertebrates and are widely used as biomaterials for a variety of applications including drug delivery and cellular and tissue engineering. In these applications, the mechanics of this hierarchically structured protein play a key role, as does its chemical composition. To facilitate investigation into how gene mutations of collagen lead to disease as well as the rational development of tunable mechanical and chemical properties of this full-length protein, production of recombinant expressed protein is required.
Here, we present a human type II procollagen expression system that produces full-length procollagen utilizing a previously characterized human fibrosarcoma cell line for production. The system exploits a non-covalently linked fluorescence readout for gene expression to facilitate screening of cell lines. Biochemical and biophysical characterization of the secreted, purified protein are used to demonstrate the proper formation and function of the protein. Assays to demonstrate fidelity include proteolytic digestion, mass spectrometric sequence and posttranslational composition analysis, circular dichroism spectroscopy, single-molecule stretching with optical tweezers, atomic-force microscopy imaging of fibril assembly, and transmission electron microscopy imaging of self-assembled fibrils.
Using a mammalian expression system, we produced full-length recombinant human type II procollagen. The integrity of the collagen preparation was verified by various structural and degradation assays. This system provides a platform from which to explore new directions in collagen manipulation.
DOI
Triple helical collagens are the most abundant structural protein in vertebrates and are widely used as biomaterials for a variety of applications including drug delivery and cellular and tissue engineering. In these applications, the mechanics of this hierarchically structured protein play a key role, as does its chemical composition. To facilitate investigation into how gene mutations of collagen lead to disease as well as the rational development of tunable mechanical and chemical properties of this full-length protein, production of recombinant expressed protein is required.
Here, we present a human type II procollagen expression system that produces full-length procollagen utilizing a previously characterized human fibrosarcoma cell line for production. The system exploits a non-covalently linked fluorescence readout for gene expression to facilitate screening of cell lines. Biochemical and biophysical characterization of the secreted, purified protein are used to demonstrate the proper formation and function of the protein. Assays to demonstrate fidelity include proteolytic digestion, mass spectrometric sequence and posttranslational composition analysis, circular dichroism spectroscopy, single-molecule stretching with optical tweezers, atomic-force microscopy imaging of fibril assembly, and transmission electron microscopy imaging of self-assembled fibrils.
Using a mammalian expression system, we produced full-length recombinant human type II procollagen. The integrity of the collagen preparation was verified by various structural and degradation assays. This system provides a platform from which to explore new directions in collagen manipulation.
DOI
Single optical tweezers based on elliptical core fiber
Yu Zhang, Li Zhao, Yunhao Chen, Zhihai Liu, Yaxun Zhang, Enming Zhao, Jun Yang, Libo Yuan
We propose and demonstrate a new single optical tweezers based on an elliptical core fiber, which can realize the trapped yeast cell rotation with a precise and simple control. Due to the elliptical shape of the fiber core, the LP11 mode beam can propagate stably. When we rotate the fiber tip, the LP11 mode beam will also rotate along with the fiber tip, which helps to realize the trapped micro-particle rotation. By using this method, we can easily realize the rotation of the trapped yeast cells, the rotating angle of the yeast cell is same as the elliptical core fiber tip.
DOI
We propose and demonstrate a new single optical tweezers based on an elliptical core fiber, which can realize the trapped yeast cell rotation with a precise and simple control. Due to the elliptical shape of the fiber core, the LP11 mode beam can propagate stably. When we rotate the fiber tip, the LP11 mode beam will also rotate along with the fiber tip, which helps to realize the trapped micro-particle rotation. By using this method, we can easily realize the rotation of the trapped yeast cells, the rotating angle of the yeast cell is same as the elliptical core fiber tip.
DOI
Thursday, December 17, 2015
Optically Induced Forces Imposed in an Optical Funnel on a Stream of Particles in Air or Vacuum
Niko Eckerskorn, Richard Bowman, Richard A. Kirian, Salah Awel, Max Wiedorn, Jochen Küpper, Miles J. Padgett, Henry N. Chapman, and Andrei V. Rode
Optical trapping of light-absorbing particles in a gaseous environment is governed by a laser-induced photophoretic force, which can be orders of magnitude stronger than the force of radiation pressure induced by the same light intensity. In spite of many experimental studies, the exact theoretical background underlying the photophoretic force and the prediction of its influence on the particle motion is still in its infancy. Here, we report the results of a quantitative analysis of the photophoretic force and the stiffness of trapping achieved by levitating graphite and graphite-coated glass shells of calibrated sizes in an upright diverging hollow-core vortex beam, which we refer to as an “optical funnel”. The measurements of forces are conducted in air at various gas pressures in the range from 5 mbar to 2 bar. The results of these measurements lay the foundation for mapping the optically induced force to the intensity distribution in the trap. The mapping, in turn, provides the necessary information to model flight trajectories of particles of various sizes entering the beam at given initial speed and position relative to the beam axis. Finally, we determine the limits of the parameter space for the particle speed, size, and radial offset to the beam axis, all linked to the laser power and the particular laser-beam structure. These results establish the grounds for developing a touch-free optical system for precisely positioning submicrometer bioparticles at the focal spot of an x-ray free-electron laser, which will significantly enhance the efficiency of studying nanoscale morphology of proteins and biomolecules in femtosecond coherent diffractive imaging experiments.
DOI
Optical trapping of light-absorbing particles in a gaseous environment is governed by a laser-induced photophoretic force, which can be orders of magnitude stronger than the force of radiation pressure induced by the same light intensity. In spite of many experimental studies, the exact theoretical background underlying the photophoretic force and the prediction of its influence on the particle motion is still in its infancy. Here, we report the results of a quantitative analysis of the photophoretic force and the stiffness of trapping achieved by levitating graphite and graphite-coated glass shells of calibrated sizes in an upright diverging hollow-core vortex beam, which we refer to as an “optical funnel”. The measurements of forces are conducted in air at various gas pressures in the range from 5 mbar to 2 bar. The results of these measurements lay the foundation for mapping the optically induced force to the intensity distribution in the trap. The mapping, in turn, provides the necessary information to model flight trajectories of particles of various sizes entering the beam at given initial speed and position relative to the beam axis. Finally, we determine the limits of the parameter space for the particle speed, size, and radial offset to the beam axis, all linked to the laser power and the particular laser-beam structure. These results establish the grounds for developing a touch-free optical system for precisely positioning submicrometer bioparticles at the focal spot of an x-ray free-electron laser, which will significantly enhance the efficiency of studying nanoscale morphology of proteins and biomolecules in femtosecond coherent diffractive imaging experiments.
DOI
Optical Trapping of Nanoparticles on a Silicon Subwavelength Grating and Their Detection by an Ellipsometric Technique
Naoya Taki, Yasuhiro Mizutan, Tetsuo Iwata, Takao Kojima, Hiroki Yamamoto & Takahiro Kozawa
A method and setup are proposed for trapping and detecting nanoparticles dispersed in a nanocomposite solution using periodically localized light generated by a subwavelength transmission grating. By numerical simulations, it is shown that there is an optimum duty ratio of the grating to produce the periodically localized light. Experimental results are presented for Au/γ-Fe2O3 composite nanoparticles having a diameter of 21.0 nm trapped on a silicon subwavelength rectangular grating and detected ellipsometrically. The technique should prove useful for evaluating optical and mechanical properties of nanocomposite materials.
DOI
A method and setup are proposed for trapping and detecting nanoparticles dispersed in a nanocomposite solution using periodically localized light generated by a subwavelength transmission grating. By numerical simulations, it is shown that there is an optimum duty ratio of the grating to produce the periodically localized light. Experimental results are presented for Au/γ-Fe2O3 composite nanoparticles having a diameter of 21.0 nm trapped on a silicon subwavelength rectangular grating and detected ellipsometrically. The technique should prove useful for evaluating optical and mechanical properties of nanocomposite materials.
DOI
Wednesday, December 16, 2015
Constructing 3D microtubule networks using holographic optical trapping
J. Bergman, O. Osunbayo & M. Vershinin
Developing abilities to assemble nanoscale structures is a major scientific and engineering challenge. We report a technique which allows precise positioning and manipulation of individual rigid filaments, enabling construction of custom-designed 3D filament networks. This approach uses holographic optical trapping (HOT) for nano-positioning and microtubules (MTs) as network building blocks. MTs are desirable engineering components due to their high aspect ratio, rigidity, and their ability to serve as substrate for directed nano-transport, reflecting their roles in the eukaryotic cytoskeleton. The 3D architecture of MT cytoskeleton is a significant component of its function, however experimental tools to study the roles of this geometric complexity in a controlled environment have been lacking. We demonstrate the broad capabilities of our system by building a self-supporting 3D MT-based nanostructure and by conducting a MT-based transport experiment on a dynamically adjustable 3D MT intersection. Our methodology not only will advance studies of cytoskeletal networks (and associated processes such as MT-based transport) but will also likely find use in engineering nanostructures and devices.
DOI
Developing abilities to assemble nanoscale structures is a major scientific and engineering challenge. We report a technique which allows precise positioning and manipulation of individual rigid filaments, enabling construction of custom-designed 3D filament networks. This approach uses holographic optical trapping (HOT) for nano-positioning and microtubules (MTs) as network building blocks. MTs are desirable engineering components due to their high aspect ratio, rigidity, and their ability to serve as substrate for directed nano-transport, reflecting their roles in the eukaryotic cytoskeleton. The 3D architecture of MT cytoskeleton is a significant component of its function, however experimental tools to study the roles of this geometric complexity in a controlled environment have been lacking. We demonstrate the broad capabilities of our system by building a self-supporting 3D MT-based nanostructure and by conducting a MT-based transport experiment on a dynamically adjustable 3D MT intersection. Our methodology not only will advance studies of cytoskeletal networks (and associated processes such as MT-based transport) but will also likely find use in engineering nanostructures and devices.
DOI
Plasmonic absorption activated trapping and assembling of colloidal crystals with non-resonant continuous gold films
Zhiwen Kang, Jiajie Chen, Shu-Yuen Wu and Ho-Pui Ho
Here we report the realization of trapping and assembly of colloidal crystals on continuous gold thin films based on the combined effect of thermophoresis and thermal convection associated with plasmonic optical heating. In the system, the stabilized trapping phenomenon is driven by thermophoretic forces caused by a temperature gradient which pushes the target particles from cold to hot regions and always in an opposite direction to the axial convective drag forces. Furthermore, the lateral convective flow of an aqueous medium accelerates the formation of the trap considerably by dragging target particles into the hot region from a long distance. The influence of salt concentration on the trapping behavior has also been investigated. Typically the threshold optical power density is in the order of microwatts per square micrometer (∼μW μm−2). We anticipate that the reported optical trapping approach may find many potential applications in biophysics, life sciences, and lab-on-a-chip devices.
DOI
Here we report the realization of trapping and assembly of colloidal crystals on continuous gold thin films based on the combined effect of thermophoresis and thermal convection associated with plasmonic optical heating. In the system, the stabilized trapping phenomenon is driven by thermophoretic forces caused by a temperature gradient which pushes the target particles from cold to hot regions and always in an opposite direction to the axial convective drag forces. Furthermore, the lateral convective flow of an aqueous medium accelerates the formation of the trap considerably by dragging target particles into the hot region from a long distance. The influence of salt concentration on the trapping behavior has also been investigated. Typically the threshold optical power density is in the order of microwatts per square micrometer (∼μW μm−2). We anticipate that the reported optical trapping approach may find many potential applications in biophysics, life sciences, and lab-on-a-chip devices.
DOI
Zebrafish as a model system for characterization of nanoparticles against cancer
Lasse Evensen, Patrick L. Johansen, Gerbrand Koster, Kaizheng Zhu, Lars Herfindal, Martin Speth, Federico Fenaroli, Jon Hildahl, Shahla Bagherifam, Claudia Tulotta, Lina Prasmickaite, Gunhild M. Mælandsmo, Ewa Snaar-Jagalska and Gareth Griffiths
Therapeutic nanoparticles (NPs) have great potential to deliver drugs against human diseases. Encapsulation of drugs in NPs protects them from being metabolized, while they are delivered specifically to a target site, thereby reducing toxicity and other side-effects. However, non-specific tissue accumulation of NPs, for example in macrophages, especially in the spleen and liver is a general problem with many NPs being developed for cancer therapy. To address the problem of non-specific tissue accumulation of NPs we describe the development of the zebrafish embryo as a transparent vertebrate system for characterization of NPs against cancer. We show that injection of human cancer cells results in tumor-like structures, and that subsequently injected fluorescent NPs, either made of polystyrene or liposomes can be imaged in real-time. NP biodistribution and general in vivo properties can be easily monitored in embryos having selective fluorescent labeling of specific tissues. We demonstrate in vitro, by using optical tweezer micromanipulation, microscopy and flow cytometry that polyethylene glycol (PEG) coating of NPs decreases the level of adhesion of NPs to macrophages, and also to cancer cells. In vivo in zebrafish embryos, PEG coating resulted in longer NP circulation times, decreased macrophage uptake, and reduced adhesion to the endothelium. Importantly, liposomes were observed to accumulate passively and selectively in tumor-like structures comprised of human cancer cells. These results show that zebrafish embryo is a powerful system for microscopy-based screening of NPs on the route to preclinical testing.
DOI
Therapeutic nanoparticles (NPs) have great potential to deliver drugs against human diseases. Encapsulation of drugs in NPs protects them from being metabolized, while they are delivered specifically to a target site, thereby reducing toxicity and other side-effects. However, non-specific tissue accumulation of NPs, for example in macrophages, especially in the spleen and liver is a general problem with many NPs being developed for cancer therapy. To address the problem of non-specific tissue accumulation of NPs we describe the development of the zebrafish embryo as a transparent vertebrate system for characterization of NPs against cancer. We show that injection of human cancer cells results in tumor-like structures, and that subsequently injected fluorescent NPs, either made of polystyrene or liposomes can be imaged in real-time. NP biodistribution and general in vivo properties can be easily monitored in embryos having selective fluorescent labeling of specific tissues. We demonstrate in vitro, by using optical tweezer micromanipulation, microscopy and flow cytometry that polyethylene glycol (PEG) coating of NPs decreases the level of adhesion of NPs to macrophages, and also to cancer cells. In vivo in zebrafish embryos, PEG coating resulted in longer NP circulation times, decreased macrophage uptake, and reduced adhesion to the endothelium. Importantly, liposomes were observed to accumulate passively and selectively in tumor-like structures comprised of human cancer cells. These results show that zebrafish embryo is a powerful system for microscopy-based screening of NPs on the route to preclinical testing.
DOI
Tuesday, December 15, 2015
Simple method to measure and analyze the fluctuations of a small particle in biopolymer solutions
Masafumi Kuroda and Yoshihiro Murayama
We developed a simple method to investigate the motion of a small particle in biopolymer solutions. Using optical tweezers with low stiffness, a trapped probe particle fluctuates widely for a long time along the light axis, which reflects the rheological properties of the surrounding environment. We present a convenient technique for three-dimensional position tracking and the analysis focused on the distribution of particle positions and its variance in a given time interval. It allows us to obtain useful information about the dynamics of a small particle in a wide range from a free diffusive motion to a constrained motion with statistical significance. We applied this method to investigate the dynamics in collagen and DNA solutions; it was found that a collagen solution behaves as a simple viscous liquid and a DNA solution has apparent elasticity due to the slow relaxation of the configuration of molecules.
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DOI
Effect of elastic colored noise in the hopping dynamics of single molecules in stretching experiments
M. Hidalgo-Soria, A. Pérez-Madrid, and I. Santamaría-Holek
The influence of colored noise induced by elastic fluctuations in single-molecule stretching experiments is theoretically and numerically studied. Unlike in the thermal white noise case currently considered in the literature, elastically induced hopping dynamics between folded and unfolded states is manifested through critical oscillations showing smaller end-to-end distance fluctuations (δx∼1.25nm) within the free energy wells corresponding to both states. Our results are derived by analyzing the elastic coupling between the Handle-Molecule-Handle system and the laser optical tweezers (LOT) array. It is shown that an Ornstein-Uhlenbeck process related to this elastic coupling may trigger the hopping transitions via a colored noise with an intensity proportional to the elastic constant of the LOT array. Evolution equations of the variables of the system were derived by using the irreversible thermodynamics of small systems recently proposed. Theoretical expressions for the corresponding stationary probability densities are provided and the viability of inferring the shape of the free energy from direct measurements is discussed.
DOI
The influence of colored noise induced by elastic fluctuations in single-molecule stretching experiments is theoretically and numerically studied. Unlike in the thermal white noise case currently considered in the literature, elastically induced hopping dynamics between folded and unfolded states is manifested through critical oscillations showing smaller end-to-end distance fluctuations (δx∼1.25nm) within the free energy wells corresponding to both states. Our results are derived by analyzing the elastic coupling between the Handle-Molecule-Handle system and the laser optical tweezers (LOT) array. It is shown that an Ornstein-Uhlenbeck process related to this elastic coupling may trigger the hopping transitions via a colored noise with an intensity proportional to the elastic constant of the LOT array. Evolution equations of the variables of the system were derived by using the irreversible thermodynamics of small systems recently proposed. Theoretical expressions for the corresponding stationary probability densities are provided and the viability of inferring the shape of the free energy from direct measurements is discussed.
DOI
Cellobiohydrolase 1 from Trichoderma reesei degrades cellulose in single cellobiose steps
Sonia K. Brady, Sarangapani Sreelatha, Yinnian Feng, Shishir P. S. Chundawat & Matthew J Lang
Cellobiohydrolase 1 from Trichoderma reesei (TrCel7A) processively hydrolyses cellulose into cellobiose. Although enzymatic techniques have been established as promising tools in biofuel production, a clear understanding of the motor’s mechanistic action has yet to be revealed. Here, we develop an optical tweezers-based single-molecule (SM) motility assay for precision tracking of TrCel7A. Direct observation of motility during degradation reveals processive runs and distinct steps on the scale of 1 nm. Our studies suggest TrCel7A is not mechanically limited, can work against 20 pN loads and speeds up when assisted. Temperature-dependent kinetic studies establish the energy requirements for the fundamental stepping cycle, which likely includes energy from glycosidic bonds and other sources. Through SM measurements of isolated TrCel7A domains, we determine that the catalytic domain alone is sufficient for processive motion, providing insight into TrCel7A’s molecular motility mechanism.
DOI
Cellobiohydrolase 1 from Trichoderma reesei (TrCel7A) processively hydrolyses cellulose into cellobiose. Although enzymatic techniques have been established as promising tools in biofuel production, a clear understanding of the motor’s mechanistic action has yet to be revealed. Here, we develop an optical tweezers-based single-molecule (SM) motility assay for precision tracking of TrCel7A. Direct observation of motility during degradation reveals processive runs and distinct steps on the scale of 1 nm. Our studies suggest TrCel7A is not mechanically limited, can work against 20 pN loads and speeds up when assisted. Temperature-dependent kinetic studies establish the energy requirements for the fundamental stepping cycle, which likely includes energy from glycosidic bonds and other sources. Through SM measurements of isolated TrCel7A domains, we determine that the catalytic domain alone is sufficient for processive motion, providing insight into TrCel7A’s molecular motility mechanism.
DOI
Fiber-Based, Injection-Molded Optofluidic Systems: Improvements in Assembly and Applications
Marco Matteucci, Marco Triches, Giovanni Nava, Anders Kristensen, Mark R. Pollard, Kirstine Berg-Sørensen and Rafael J. Taboryski
We present a method to fabricate polymer optofluidic systems by means of injection molding that allow the insertion of standard optical fibers. The chip fabrication and assembly methods produce large numbers of robust optofluidic systems that can be easily assembled and disposed of, yet allow precise optical alignment and improve delivery of optical power. Using a multi-level chip fabrication process, complex channel designs with extremely vertical sidewalls, and dimensions that range from few tens of nanometers to hundreds of microns can be obtained. The technology has been used to align optical fibers in a quick and precise manner, with a lateral alignment accuracy of 2.7 ± 1.8 μm. We report the production, assembly methods, and the characterization of the resulting injection-molded chips for Lab-on-Chip (LoC) applications. We demonstrate the versatility of this technology by carrying out two types of experiments that benefit from the improved optical system: optical stretching of red blood cells (RBCs) and Raman spectroscopy of a solution loaded into a hollow core fiber. The advantages offered by the presented technology are intended to encourage the use of LoC technology for commercialization and educational purposes.
DOI
We present a method to fabricate polymer optofluidic systems by means of injection molding that allow the insertion of standard optical fibers. The chip fabrication and assembly methods produce large numbers of robust optofluidic systems that can be easily assembled and disposed of, yet allow precise optical alignment and improve delivery of optical power. Using a multi-level chip fabrication process, complex channel designs with extremely vertical sidewalls, and dimensions that range from few tens of nanometers to hundreds of microns can be obtained. The technology has been used to align optical fibers in a quick and precise manner, with a lateral alignment accuracy of 2.7 ± 1.8 μm. We report the production, assembly methods, and the characterization of the resulting injection-molded chips for Lab-on-Chip (LoC) applications. We demonstrate the versatility of this technology by carrying out two types of experiments that benefit from the improved optical system: optical stretching of red blood cells (RBCs) and Raman spectroscopy of a solution loaded into a hollow core fiber. The advantages offered by the presented technology are intended to encourage the use of LoC technology for commercialization and educational purposes.
DOI
Monday, December 14, 2015
Single exosome study reveals subpopulations distributed among cell lines with variability related to membrane content
Zachary J. Smith, Changwon Lee, Tatu Rojalin, Randy P. Carney, Sidhartha Hazari, Alisha Knudson, Kit Lam, Heikki Saari, Elisa Lazaro Ibañez, Tapani Viitala, Timo Laaksonen, Marjo Yliperttula and Sebastian Wachsmann-Hogiu
Current analysis of exosomes focuses primarily on bulk analysis, where exosome-to-exosome variability cannot be assessed. In this study, we used Raman spectroscopy to study the chemical composition of single exosomes. We measured spectra of individual exosomes from 8 cell lines. Cell-line-averaged spectra varied considerably, reflecting the variation in total exosomal protein, lipid, genetic, and cytosolic content. Unexpectedly, single exosomes isolated from the same cell type also exhibited high spectral variability. Subsequent spectral analysis revealed clustering of single exosomes into 4 distinct groups that were not cell-line specific. Each group contained exosomes from multiple cell lines, and most cell lines had exosomes in multiple groups. The differences between these groups are related to chemical differences primarily due to differing membrane composition. Through a principal components analysis, we identified that the major sources of spectral variation among the exosomes were in cholesterol content, relative expression of phospholipids to cholesterol, and surface protein expression. For example, exosomes derived from cancerous versus non-cancerous cell lines can be largely separated based on their relative expression of cholesterol and phospholipids. We are the first to indicate that exosome subpopulations are shared among cell types, suggesting distributed exosome functionality. The origins of these differences are likely related to the specific role of extracellular vesicle subpopulations in both normal cell function and carcinogenesis, and they may provide diagnostic potential at the single exosome level.
DOI
Current analysis of exosomes focuses primarily on bulk analysis, where exosome-to-exosome variability cannot be assessed. In this study, we used Raman spectroscopy to study the chemical composition of single exosomes. We measured spectra of individual exosomes from 8 cell lines. Cell-line-averaged spectra varied considerably, reflecting the variation in total exosomal protein, lipid, genetic, and cytosolic content. Unexpectedly, single exosomes isolated from the same cell type also exhibited high spectral variability. Subsequent spectral analysis revealed clustering of single exosomes into 4 distinct groups that were not cell-line specific. Each group contained exosomes from multiple cell lines, and most cell lines had exosomes in multiple groups. The differences between these groups are related to chemical differences primarily due to differing membrane composition. Through a principal components analysis, we identified that the major sources of spectral variation among the exosomes were in cholesterol content, relative expression of phospholipids to cholesterol, and surface protein expression. For example, exosomes derived from cancerous versus non-cancerous cell lines can be largely separated based on their relative expression of cholesterol and phospholipids. We are the first to indicate that exosome subpopulations are shared among cell types, suggesting distributed exosome functionality. The origins of these differences are likely related to the specific role of extracellular vesicle subpopulations in both normal cell function and carcinogenesis, and they may provide diagnostic potential at the single exosome level.
DOI
Surface-modified complex SU-8 microstructures for indirect optical manipulation of single cells
Badri L. Aekbote, Tamás Fekete, Jaroslaw Jacak, Gaszton Vizsnyiczai, Pál Ormos, and Lóránd Kelemen
We introduce a method that combines two-photon polymerization (TPP) and surface functionalization to enable the indirect optical manipulation of live cells. TPP-made 3D microstructures were coated specifically with a multilayer of the protein streptavidin and non-specifically with IgG antibody using polyethylene glycol diamine as a linker molecule. Protein density on their surfaces was quantified for various coating methods. The streptavidin-coated structures were shown to attach to biotinated cells reproducibly. We performed basic indirect optical micromanipulation tasks with attached structure-cell couples using complex structures and a multi-focus optical trap. The use of such extended manipulators for indirect optical trapping ensures to keep a safe distance between the trapping beams and the sensitive cell and enables their 6 degrees of freedom actuation.
DOI
We introduce a method that combines two-photon polymerization (TPP) and surface functionalization to enable the indirect optical manipulation of live cells. TPP-made 3D microstructures were coated specifically with a multilayer of the protein streptavidin and non-specifically with IgG antibody using polyethylene glycol diamine as a linker molecule. Protein density on their surfaces was quantified for various coating methods. The streptavidin-coated structures were shown to attach to biotinated cells reproducibly. We performed basic indirect optical micromanipulation tasks with attached structure-cell couples using complex structures and a multi-focus optical trap. The use of such extended manipulators for indirect optical trapping ensures to keep a safe distance between the trapping beams and the sensitive cell and enables their 6 degrees of freedom actuation.
DOI
Photoinduced magnetic force between nanostructures
Caner Guclu, Venkata Ananth Tamma, Hemantha Kumar Wickramasinghe, and Filippo Capolino
Photoinduced magnetic force between nanostructures, at optical frequencies, is investigated theoretically. Till now optical magnetic effects were not used in scanning probe microscopy because of the vanishing natural magnetism with increasing frequency. On the other hand, artificial magnetism in engineered nanostructures led to the development of measurable optical magnetism. Here two examples of nanoprobes that are able to generate strong magnetic dipolar fields at optical frequency are investigated: first, an ideal magnetically polarizable nanosphere and then a circular cluster of silver nanospheres that has a looplike collective plasmonic resonance equivalent to a magnetic dipole. Magnetic forces are evaluated based on nanostructure polarizabilities, i.e., induced magnetic dipoles, and magnetic-near field evaluations. As an initial assessment on the possibility of a magnetic nanoprobe to detect magnetic forces, we consider two identical magnetically polarizable nanoprobes and observe magnetic forces on the order of piconewtons, thereby bringing it within detection limits of conventional atomic force microscopes at ambient pressure and temperature. The detection of magnetic force is a promising method in studying optical magnetic transitions that can be the basis of innovative spectroscopy applications.
DOI
Photoinduced magnetic force between nanostructures, at optical frequencies, is investigated theoretically. Till now optical magnetic effects were not used in scanning probe microscopy because of the vanishing natural magnetism with increasing frequency. On the other hand, artificial magnetism in engineered nanostructures led to the development of measurable optical magnetism. Here two examples of nanoprobes that are able to generate strong magnetic dipolar fields at optical frequency are investigated: first, an ideal magnetically polarizable nanosphere and then a circular cluster of silver nanospheres that has a looplike collective plasmonic resonance equivalent to a magnetic dipole. Magnetic forces are evaluated based on nanostructure polarizabilities, i.e., induced magnetic dipoles, and magnetic-near field evaluations. As an initial assessment on the possibility of a magnetic nanoprobe to detect magnetic forces, we consider two identical magnetically polarizable nanoprobes and observe magnetic forces on the order of piconewtons, thereby bringing it within detection limits of conventional atomic force microscopes at ambient pressure and temperature. The detection of magnetic force is a promising method in studying optical magnetic transitions that can be the basis of innovative spectroscopy applications.
DOI
Friday, December 11, 2015
Cellular Temperature Measurement by Dielectrophoretic Impedance Measurement Method
Ryo KIDO, Kozo TAGUCHI
Dielectrophoretic impedance measurement (DEPIM) method has recently attracted attention because the method is simple and immediately. We demonstrated that the impedance between the electrodes with trapped cell has thermal property and measured cellular temperature by the impedance change even if the number of trapped cells did not change. However, this result was not observed when cells were not trapped between the electrodes.
In summary, the cellular temperature could be measured by observing the change in shunt voltage. In this study, we developed cellular temperature measurement system and it was expected to be used in biosensor application. In the future, we want to measure single cellular temperature with isolation by optical tweezers.
DOI
Dielectrophoretic impedance measurement (DEPIM) method has recently attracted attention because the method is simple and immediately. We demonstrated that the impedance between the electrodes with trapped cell has thermal property and measured cellular temperature by the impedance change even if the number of trapped cells did not change. However, this result was not observed when cells were not trapped between the electrodes.
In summary, the cellular temperature could be measured by observing the change in shunt voltage. In this study, we developed cellular temperature measurement system and it was expected to be used in biosensor application. In the future, we want to measure single cellular temperature with isolation by optical tweezers.
DOI
Femtosecond Fiber Laser Applying for Cell Fusion
Trang Dang NGUYEN, Yoshihiro MIZUTA, Kozo TAGUCHI
We developed an actively mode-locked fiber laser that can generate 295 fs pulses at 9.188 MHz repetition rate. We built up a laser-induced cell fusion, in which the developed femtosecond laser was used as the laser source for both optical tweezers mode and laser scalpel mode, and thus improving cost-effectiveness. The cell fusion system also used a transparent dielectrophoresis chip as the specimen stage to create and manipulate the pearl chain of two or multiple cells for facilitating the cell fusion processes. We successfully developed the first optical tweezers using femtosecond fiber laser operating at 1530 nm, which can trap and transport cells effectively. With this developed system, we obtained the laser-induced fusion of red cabbage protoplasts. We also proposed a experimental cell fusion procedure which allows precisely selective cell fusion at the single-cell level. Therefore, the developed system would benefit basic research in biotechnology and biomedicine.
DOI
We developed an actively mode-locked fiber laser that can generate 295 fs pulses at 9.188 MHz repetition rate. We built up a laser-induced cell fusion, in which the developed femtosecond laser was used as the laser source for both optical tweezers mode and laser scalpel mode, and thus improving cost-effectiveness. The cell fusion system also used a transparent dielectrophoresis chip as the specimen stage to create and manipulate the pearl chain of two or multiple cells for facilitating the cell fusion processes. We successfully developed the first optical tweezers using femtosecond fiber laser operating at 1530 nm, which can trap and transport cells effectively. With this developed system, we obtained the laser-induced fusion of red cabbage protoplasts. We also proposed a experimental cell fusion procedure which allows precisely selective cell fusion at the single-cell level. Therefore, the developed system would benefit basic research in biotechnology and biomedicine.
DOI
Metal Coated Chemically Etched Fiber Probe for Single Cell Manipulation and Isolation
Kozo TAGUCHI, Ryo KIDO, Yoshihiro MIZUTA
In this paper, dielectrophoresis tweezers using metal coated chemically etched fiber was proposed for cell manipulation and isolation. We proposed a simple and low cost dielectrophoretic device for picking out and relocating single target cells. The device consists of metal coated chemically etched fibers and an AC signal generator. It does not require microfabrication technologies or sophisticated electronics. From experimental results, it was found that our proposed dielectrophoretic manipulator could discriminate between live and dead cells. We also could see the cell reproduction of yeast cells trapped and isolated using our proposed dielectrophoresis tweezers.
In this paper, dielectrophoresis tweezers using metal coated chemically etched fiber was proposed for cell manipulation and isolation. We proposed a simple and low cost dielectrophoretic device for picking out and relocating single target cells. The device consists of metal coated chemically etched fibers and an AC signal generator. It does not require microfabrication technologies or sophisticated electronics. From experimental results, it was found that our proposed dielectrophoretic manipulator could discriminate between live and dead cells. We also could see the cell reproduction of yeast cells trapped and isolated using our proposed dielectrophoresis tweezers.
Lab-on-Chip System Combined Optical Tweezers and Dielectrophoresis
Yoshihiro MIZUTA, Kozo TAGUCHI
Cell manipulation and operation have played important roles in modern biotechnology, hence a number of researchers have developed them during the past decades. In this paper we introduce two techniques based on dielectrophoresis (DEP) and optical tweezers: Distinction between viable cells and non-viable cells using DEP, Manipulating cells captured by positive DEP (pDEP) using optical tweezers. And then we suggest a system combined both DEP and optical tweezers, that would be a useful tool in biotechnology. This system will be useful to perform cell operation and other applications in biotechnology.
DOI
Cell manipulation and operation have played important roles in modern biotechnology, hence a number of researchers have developed them during the past decades. In this paper we introduce two techniques based on dielectrophoresis (DEP) and optical tweezers: Distinction between viable cells and non-viable cells using DEP, Manipulating cells captured by positive DEP (pDEP) using optical tweezers. And then we suggest a system combined both DEP and optical tweezers, that would be a useful tool in biotechnology. This system will be useful to perform cell operation and other applications in biotechnology.
DOI
Thursday, December 10, 2015
Self-induced back-action optical trapping in nanophotonic systems
Lukas Neumeier, Romain Quidant and Darrick E Chang
Optical trapping is an indispensable tool in physics and the life sciences. However, there is a clear trade off between the size of a particle to be trapped, its spatial confinement, and the intensities required. This is due to the decrease in optical response of smaller particles and the diffraction limit that governs the spatial variation of optical fields. It is thus highly desirable to find techniques that surpass these bounds. Recently, a number of experiments using nanophotonic cavities have observed a qualitatively different trapping mechanism described as 'self-induced back-action trapping' (SIBA). In these systems, the particle motion couples to the resonance frequency of the cavity, which results in a strong interplay between the intra-cavity field intensity and the forces exerted. Here, we provide a theoretical description that for the first time captures the remarkable range of consequences. In particular, we show that SIBA can be exploited to yield dynamic reshaping of trap potentials, strongly sub-wavelength trap features, and significant reduction of intensities seen by the particle, which should have important implications for future trapping technologies.
DOI
Optical trapping is an indispensable tool in physics and the life sciences. However, there is a clear trade off between the size of a particle to be trapped, its spatial confinement, and the intensities required. This is due to the decrease in optical response of smaller particles and the diffraction limit that governs the spatial variation of optical fields. It is thus highly desirable to find techniques that surpass these bounds. Recently, a number of experiments using nanophotonic cavities have observed a qualitatively different trapping mechanism described as 'self-induced back-action trapping' (SIBA). In these systems, the particle motion couples to the resonance frequency of the cavity, which results in a strong interplay between the intra-cavity field intensity and the forces exerted. Here, we provide a theoretical description that for the first time captures the remarkable range of consequences. In particular, we show that SIBA can be exploited to yield dynamic reshaping of trap potentials, strongly sub-wavelength trap features, and significant reduction of intensities seen by the particle, which should have important implications for future trapping technologies.
DOI
Depletion interactions and modulation of DNA-intercalators binding: Opposite behavior of the “neutral” polymer poly(ethylene-glycol)
F. A. P. Crisafuli, L. H. M. da Silva, G. M. D. Ferreira, E. B. Ramos and M. S. Rocha
In this work we have investigated the role of high molecular weight poly(ethylene-glycol) 8000 (PEG 8000) in modulating the interactions of the DNA molecule with two hydrophobic compounds: Ethidium Bromide (EtBr) and GelRed (GR). Both compounds are DNA intercalators and are used here to mimic the behavior of more complex DNA ligands such as chemotherapeutic drugs and proteins whose domains intercalate DNA. By means of single-molecule stretching experiments, we have been able to show that PEG 8000 strongly shifts the binding equilibrium between the intercalators and the DNA even at very low concentrations (1% in mass). Additionally, microcalorimetry experiments were performed to estimate the strength of the interaction between PEG and the DNA ligands. Our results suggest that PEG, depending on the system under study, may act as an “inert polymer” with no enthalpic contribution in some processes but, on the other hand, it may as well be an active (non-neutral) osmolyte in the context of modulating the activity of the reactants and products involved in DNA-ligand interactions.
DOI
In this work we have investigated the role of high molecular weight poly(ethylene-glycol) 8000 (PEG 8000) in modulating the interactions of the DNA molecule with two hydrophobic compounds: Ethidium Bromide (EtBr) and GelRed (GR). Both compounds are DNA intercalators and are used here to mimic the behavior of more complex DNA ligands such as chemotherapeutic drugs and proteins whose domains intercalate DNA. By means of single-molecule stretching experiments, we have been able to show that PEG 8000 strongly shifts the binding equilibrium between the intercalators and the DNA even at very low concentrations (1% in mass). Additionally, microcalorimetry experiments were performed to estimate the strength of the interaction between PEG and the DNA ligands. Our results suggest that PEG, depending on the system under study, may act as an “inert polymer” with no enthalpic contribution in some processes but, on the other hand, it may as well be an active (non-neutral) osmolyte in the context of modulating the activity of the reactants and products involved in DNA-ligand interactions.
DOI
Wednesday, December 9, 2015
Transformation and patterning of supermicelles using dynamic holographic assembly
Oliver E.C. Gould, Huibin Qiu, David J. Lunn, John Rowden, Robert L. Harniman, Zachary M. Hudson, Mitchell A. Winnik, Mervyn J. Miles & Ian Manners
Although the solution self-assembly of block copolymers has enabled the fabrication of a broad range of complex, functional nanostructures, their precise manipulation and patterning remain a key challenge. Here we demonstrate that spherical and linear supermicelles, supramolecular structures held together by non-covalent solvophobic and coordination interactions and formed by the hierarchical self-assembly of block copolymer micelle and block comicelle precursors, can be manipulated, transformed and patterned with mediation by dynamic holographic assembly (optical tweezers). This allows the creation of new and stable soft-matter superstructures far from equilibrium. For example, individual spherical supermicelles can be optically held in close proximity and photocrosslinked through controlled coronal chemistry to generate linear oligomeric arrays. The use of optical tweezers also enables the directed deposition and immobilization of supermicelles on surfaces, allowing the precise creation of arrays of soft-matter nano-objects with potentially diverse functionality and a range of applications.
DOI
Although the solution self-assembly of block copolymers has enabled the fabrication of a broad range of complex, functional nanostructures, their precise manipulation and patterning remain a key challenge. Here we demonstrate that spherical and linear supermicelles, supramolecular structures held together by non-covalent solvophobic and coordination interactions and formed by the hierarchical self-assembly of block copolymer micelle and block comicelle precursors, can be manipulated, transformed and patterned with mediation by dynamic holographic assembly (optical tweezers). This allows the creation of new and stable soft-matter superstructures far from equilibrium. For example, individual spherical supermicelles can be optically held in close proximity and photocrosslinked through controlled coronal chemistry to generate linear oligomeric arrays. The use of optical tweezers also enables the directed deposition and immobilization of supermicelles on surfaces, allowing the precise creation of arrays of soft-matter nano-objects with potentially diverse functionality and a range of applications.
DOI
Probing the Evaporation Dynamics of Ethanol/Gasoline Biofuel Blends Using Single Droplet Manipulation Techniques
Stella Corsetti, Rachael E.H. Miles, Craig McDonald, Yuri Belotti, Jonathan Philip Reid, Johannes Kiefer, and David McGloin
Using blends of bio-ethanol and gasoline as automotive fuel leads to a net decrease in the production of harmful emission compared to the use of pure fossil fuel. However, fuel droplet evaporation dynamics change depending on the mixing ratio. Here we use single particle manipulation techniques to study the evaporation dynamics of ethanol/gasoline blend micro-droplets. The use of an electrodynamic balance (EDB) enables measurements of the evaporation of individual droplets in a controlled environment, while optical tweezers facilitate studies of the behavior of droplets inside a spray. Hence, the combination of both methods is perfectly suited to obtain a complete picture of the evaporation process. The influence of adding varied amounts of ethanol to gasoline is investigated, and we observe that droplets with a greater fraction of ethanol take longer to evaporate. Furthermore, we find that our methods are sensitive enough to observe the presence of trace amounts of water in the droplets. A theoretical model, predicting the evaporation of ethanol and gasoline droplets in dry nitrogen gas, is used to explain the experimental results. Also a theoretical estimation of the saturation of the environment, with other aerosols, in the tweezers is carried out.
DOI
Using blends of bio-ethanol and gasoline as automotive fuel leads to a net decrease in the production of harmful emission compared to the use of pure fossil fuel. However, fuel droplet evaporation dynamics change depending on the mixing ratio. Here we use single particle manipulation techniques to study the evaporation dynamics of ethanol/gasoline blend micro-droplets. The use of an electrodynamic balance (EDB) enables measurements of the evaporation of individual droplets in a controlled environment, while optical tweezers facilitate studies of the behavior of droplets inside a spray. Hence, the combination of both methods is perfectly suited to obtain a complete picture of the evaporation process. The influence of adding varied amounts of ethanol to gasoline is investigated, and we observe that droplets with a greater fraction of ethanol take longer to evaporate. Furthermore, we find that our methods are sensitive enough to observe the presence of trace amounts of water in the droplets. A theoretical model, predicting the evaporation of ethanol and gasoline droplets in dry nitrogen gas, is used to explain the experimental results. Also a theoretical estimation of the saturation of the environment, with other aerosols, in the tweezers is carried out.
DOI
Broadband boundary effects on Brownian motion
Jianyong Mo, Akarsh Simha, and Mark G. Raizen
Brownian motion of particles in confined fluids is important for many applications, yet the effects of the boundary over a wide range of time scales are still not well understood. We report high-bandwidth, comprehensive measurements of Brownian motion of an optically trapped micrometer-sized silica sphere in water near an approximately flat wall. At short distances we observe anisotropic Brownian motion with respect to the wall. We find that surface confinement not only occurs in the long time scale diffusive regime but also in the short time scale ballistic regime, and the velocity autocorrelation function of the Brownian particle decays faster than that of a particle in bulk fluid. Furthermore, at low frequencies the thermal force loses its color due to the reflected flow from the no-slip boundary. The power spectrum of the thermal force on the particle near a no-slip boundary becomes flat at low frequencies. This detailed understanding of boundary effects on Brownian motion opens a door to developing a 3D microscope using particles as remote sensors.
DOI
Brownian motion of particles in confined fluids is important for many applications, yet the effects of the boundary over a wide range of time scales are still not well understood. We report high-bandwidth, comprehensive measurements of Brownian motion of an optically trapped micrometer-sized silica sphere in water near an approximately flat wall. At short distances we observe anisotropic Brownian motion with respect to the wall. We find that surface confinement not only occurs in the long time scale diffusive regime but also in the short time scale ballistic regime, and the velocity autocorrelation function of the Brownian particle decays faster than that of a particle in bulk fluid. Furthermore, at low frequencies the thermal force loses its color due to the reflected flow from the no-slip boundary. The power spectrum of the thermal force on the particle near a no-slip boundary becomes flat at low frequencies. This detailed understanding of boundary effects on Brownian motion opens a door to developing a 3D microscope using particles as remote sensors.
DOI
Tractor beam for fully immersed multiple objects: Long distance pulling, trapping, and rotation with a single optical set-up
Md. Masudur Rahman, Ayed Al Sayem, M. R. C. Mahdy, Md. Ehsanul Haque, Rakibul Islam, S. Tanvir-ur-Rahman Chowdhury and Md. Abdul Matin
In this article, we have theoretically demonstrated the mechanism of an active tractor beam for multiple fully immersed objects with additional abilities to yielding stable long distance levitation, a controlled rotation and a desired 3D trapping. This is demonstrated with a single optical set-up by using two coaxial, or even non-coaxial, superimposed higher order monochromatic Bessel beams of reverse helical nature and different frequencies. The superimposed beams can possess periodic intensity variations both along and around the beam-axis due to a difference in longitudinal wave-numbers and beam orders, respectively. The difference in frequencies of the two laser beams makes the intensity pattern to move along and around the beam-axis in a continuous way without manual ramping of phase, which allows for bidirectional movement of completely immersed multiple particles. The condition for increasing or decreasing the dimension of binding regions is also proposed here to manipulate multiple immersed objects of different sizes under dipole approximation.
DOI
In this article, we have theoretically demonstrated the mechanism of an active tractor beam for multiple fully immersed objects with additional abilities to yielding stable long distance levitation, a controlled rotation and a desired 3D trapping. This is demonstrated with a single optical set-up by using two coaxial, or even non-coaxial, superimposed higher order monochromatic Bessel beams of reverse helical nature and different frequencies. The superimposed beams can possess periodic intensity variations both along and around the beam-axis due to a difference in longitudinal wave-numbers and beam orders, respectively. The difference in frequencies of the two laser beams makes the intensity pattern to move along and around the beam-axis in a continuous way without manual ramping of phase, which allows for bidirectional movement of completely immersed multiple particles. The condition for increasing or decreasing the dimension of binding regions is also proposed here to manipulate multiple immersed objects of different sizes under dipole approximation.
DOI
Monday, December 7, 2015
Exploded view of higher order G-quadruplex structures through click-chemistry assisted single-molecule mechanical unfolding
Sangeetha Selvam, Zhongbo Yu and Hanbin Mao
Due to the long-range nature of high-order interactions between distal components in a biomolecule, transition dynamics of tertiary structures is often too complex to profile using conventional methods. Inspired by the exploded view in mechanical drawing, here, we used laser tweezers to mechanically dissect high-order DNA structures into two constituting G-quadruplexes in the promoter of the human telomerase reverse transcriptase (hTERT) gene. Assisted with click-chemistry coupling, we sandwiched one G-quadruplex with two dsDNA handles while leaving the other unit free. Mechanical unfolding through these handles revealed transition dynamics of the targeted quadruplex in a native environment, which is named as native mechanical segmentation (NMS). Comparison between unfolding of an NMS construct and that of truncated G-quadruplex constructs revealed a quadruplex–quadruplex interaction with 2 kcal/mol stabilization energy. After mechanically targeting the two G-quadruplexes together, the same interaction was observed during the first unfolding step. The unfolding then proceeded through disrupting the weaker G-quadruplex at the 5′-end, followed by the stronger G-quadruplex at the 3′-end via various intermediates. Such a pecking order in unfolding well reflects the hierarchical nature of nucleic acid structures. With surgery-like precisions, we anticipate this NMS approach offers unprecedented perspective to decipher dynamic transitions in complex biomacromolecules.
DOI
Due to the long-range nature of high-order interactions between distal components in a biomolecule, transition dynamics of tertiary structures is often too complex to profile using conventional methods. Inspired by the exploded view in mechanical drawing, here, we used laser tweezers to mechanically dissect high-order DNA structures into two constituting G-quadruplexes in the promoter of the human telomerase reverse transcriptase (hTERT) gene. Assisted with click-chemistry coupling, we sandwiched one G-quadruplex with two dsDNA handles while leaving the other unit free. Mechanical unfolding through these handles revealed transition dynamics of the targeted quadruplex in a native environment, which is named as native mechanical segmentation (NMS). Comparison between unfolding of an NMS construct and that of truncated G-quadruplex constructs revealed a quadruplex–quadruplex interaction with 2 kcal/mol stabilization energy. After mechanically targeting the two G-quadruplexes together, the same interaction was observed during the first unfolding step. The unfolding then proceeded through disrupting the weaker G-quadruplex at the 5′-end, followed by the stronger G-quadruplex at the 3′-end via various intermediates. Such a pecking order in unfolding well reflects the hierarchical nature of nucleic acid structures. With surgery-like precisions, we anticipate this NMS approach offers unprecedented perspective to decipher dynamic transitions in complex biomacromolecules.
DOI
FACS-style detection for real-time cell viscoelastic cytometry
Aditya Kasukurti, Charles Eggleton, Sanjay A Desai and David W M Marr
Cell mechanical properties have been established as a label-free biophysical marker of cell viability and health; however, real-time methods with significant throughput for accurately and non-destructively measuring these properties remain widely unavailable. Without appropriate labels for use with fluorescence activated cell sorters (FACS), easily implemented real-time technology for tracking cell-level mechanical properties remains a current need. Employing modulated optical forces and enabled by a low-dimensional FACS-style detection method introduced here, we present a viscoelasticity cytometer (VC) capable of real-time and continuous measure. We demonstrate utility of this approach by tracking the high-frequency cell physical properties of populations of chemically-modified cells at rates of ~ 1 s-1 and explain observations within the context of a simple theoretical model.
DOI
Cell mechanical properties have been established as a label-free biophysical marker of cell viability and health; however, real-time methods with significant throughput for accurately and non-destructively measuring these properties remain widely unavailable. Without appropriate labels for use with fluorescence activated cell sorters (FACS), easily implemented real-time technology for tracking cell-level mechanical properties remains a current need. Employing modulated optical forces and enabled by a low-dimensional FACS-style detection method introduced here, we present a viscoelasticity cytometer (VC) capable of real-time and continuous measure. We demonstrate utility of this approach by tracking the high-frequency cell physical properties of populations of chemically-modified cells at rates of ~ 1 s-1 and explain observations within the context of a simple theoretical model.
DOI
Simulation and measurement of stiffness for dual beam laser trap using residual gravity method
Zhenggang Li, Yu Shen, Huizhu Hu, Bo Jia, Minqiang Tie
Dual-beam optical trap is constituted by two beams of unfocused counter-propagating laser. It has advantages such as several equilibrium points, small optical power density, large capture range and easier integration with other technologies, which optical tweezers may not achieve. Dual-beam optical trap stiffness is an important parameter to describe its mechanical properties. On the basis of quartz-substrate dual-beam optical fibre trap unit, we demonstrate a simple method for measuring stiffness. Residual gravity offset by buoyancy may provide a tiny force for the optical trap. By measuring the relations between the captured particle's axial equilibrium position and residual gravity, we may calibrate the optical trap stiffness. This method simply requires measuring the relative position of the particle in the steady state of the optical trap, and dynamic measurement of the particles location is not needed, so we can get a high position accuracy by long time measuring, thus the accuracy of the stiffness calibration is improved. In this paper, we simulated and carried out experiments to measure the stiffness of a dual-beam fibre optical trap unit based on a quartz-substrate. We simulate the stiffness of several Mie particles with different materials and sizes in dual-beam optical trap as well as the relation between the waist radius of fundamental mode Gaussian beam and stiffness. Stiffness measurement experiment is conducted using the residual gravity method. Stiffness values measured in experiments are consistent with the simulation results, which verify the validity and accuracy of the method.
DOI
Dual-beam optical trap is constituted by two beams of unfocused counter-propagating laser. It has advantages such as several equilibrium points, small optical power density, large capture range and easier integration with other technologies, which optical tweezers may not achieve. Dual-beam optical trap stiffness is an important parameter to describe its mechanical properties. On the basis of quartz-substrate dual-beam optical fibre trap unit, we demonstrate a simple method for measuring stiffness. Residual gravity offset by buoyancy may provide a tiny force for the optical trap. By measuring the relations between the captured particle's axial equilibrium position and residual gravity, we may calibrate the optical trap stiffness. This method simply requires measuring the relative position of the particle in the steady state of the optical trap, and dynamic measurement of the particles location is not needed, so we can get a high position accuracy by long time measuring, thus the accuracy of the stiffness calibration is improved. In this paper, we simulated and carried out experiments to measure the stiffness of a dual-beam fibre optical trap unit based on a quartz-substrate. We simulate the stiffness of several Mie particles with different materials and sizes in dual-beam optical trap as well as the relation between the waist radius of fundamental mode Gaussian beam and stiffness. Stiffness measurement experiment is conducted using the residual gravity method. Stiffness values measured in experiments are consistent with the simulation results, which verify the validity and accuracy of the method.
DOI
From transverse angular momentum to photonic wheels
Andrea Aiello, Peter Banzer, Martin Neugebauer & Gerd Leuchs
Scientists have known for more than a century that light possesses both linear and angular momenta along the direction of propagation. However, only recent advances in optics have led to the notion of spinning electromagnetic fields capable of carrying angular momenta transverse to the direction of motion. Such fields enable numerous applications in nano-optics, biosensing and near-field microscopy, including three-dimensional control over atoms, molecules and nanostructures, and allowing for the realization of chiral nanophotonic interfaces and plasmonic devices. Here, we report on recent developments of optics with light carrying transverse spin. We present both the underlying principles and the latest achievements, and also highlight new capabilities and future applications emerging from this young yet already advanced field of research.
DOI
Scientists have known for more than a century that light possesses both linear and angular momenta along the direction of propagation. However, only recent advances in optics have led to the notion of spinning electromagnetic fields capable of carrying angular momenta transverse to the direction of motion. Such fields enable numerous applications in nano-optics, biosensing and near-field microscopy, including three-dimensional control over atoms, molecules and nanostructures, and allowing for the realization of chiral nanophotonic interfaces and plasmonic devices. Here, we report on recent developments of optics with light carrying transverse spin. We present both the underlying principles and the latest achievements, and also highlight new capabilities and future applications emerging from this young yet already advanced field of research.
DOI
Friday, December 4, 2015
Detecting Single Gold Nanoparticles (1.8 nm) with Ultrahigh-Q Air-Mode Photonic Crystal Nanobeam Cavities
Feng Liang and Qimin Quan
The growing applications of nanoparticles in energy and healthcare demand new metrology techniques with improved sensitivity, lower sample concentration, and affordable instrument cost. Here we demonstrate the first air-mode photonic crystal nanobeam cavity with ultrahigh Q-factor (Q = 2.5 × 105) and ultrasmall mode volume (V = 0.01λ3) at telecom wavelength. The air-mode cavity has strong field localization outside of its high-index material, thus significantly improving the sensitivity to detect nanoparticles. The strong field gradient attracts the nanoparticles to its field maximum, improving the detection efficiency. Combining these advantages, we report detecting and sizing single gold nanoparticles down to 1.8 nm in diameter (equivalently single polystyrene nanoparticle of 3 nm in diameter) with significantly reduced sample concentration (∼fM) than traditional optical techniques. In addition, the air-mode ultrahigh Q, ultrasmall V photonic crystal nanobeam cavity will be a useful platform to study strong light–matter interactions, nonlinear processes, and cavity quantum electrodynamics.
DOI
The growing applications of nanoparticles in energy and healthcare demand new metrology techniques with improved sensitivity, lower sample concentration, and affordable instrument cost. Here we demonstrate the first air-mode photonic crystal nanobeam cavity with ultrahigh Q-factor (Q = 2.5 × 105) and ultrasmall mode volume (V = 0.01λ3) at telecom wavelength. The air-mode cavity has strong field localization outside of its high-index material, thus significantly improving the sensitivity to detect nanoparticles. The strong field gradient attracts the nanoparticles to its field maximum, improving the detection efficiency. Combining these advantages, we report detecting and sizing single gold nanoparticles down to 1.8 nm in diameter (equivalently single polystyrene nanoparticle of 3 nm in diameter) with significantly reduced sample concentration (∼fM) than traditional optical techniques. In addition, the air-mode ultrahigh Q, ultrasmall V photonic crystal nanobeam cavity will be a useful platform to study strong light–matter interactions, nonlinear processes, and cavity quantum electrodynamics.
DOI
Topological binding and elastic interactions of microspheres and fibres in a nematic liquid crystal
M. Nikkhou, M. Škarabot, I. Muševič
We present a detailed analysis of topological binding and elastic interactions between a long, and micrometer-diameter fiber, and a microsphere in a homogeneously aligned nematic liquid crystal. Both objects are surface treated to produce strong perpendicular anchoring of the nematic liquid crystal. We use the opto-thermal micro-quench of the laser tweezers to produce topological defects with prescribed topological charge, such as pairs of a Saturn ring and an anti-ring, hyperbolic and radial hedgehogs on a fiber, as well as zero-charge loops. We study the entanglement and topological charge interaction between the topological defects of the fiber and sphere and we observe a huge variety of different entanglement topologies and defect-mediated elastic bindings. We explain all observed phenomena with simple topological rule: like topological charges repel each other and opposite topological charges attract. These binding mechanisms not only demonstrate the fascinating topology of nematic colloids, but also open a novel route to the assembly of very complex topological networks of fibers, spheres and other objects for applications in liquid crystal photonics.
DOI
We present a detailed analysis of topological binding and elastic interactions between a long, and micrometer-diameter fiber, and a microsphere in a homogeneously aligned nematic liquid crystal. Both objects are surface treated to produce strong perpendicular anchoring of the nematic liquid crystal. We use the opto-thermal micro-quench of the laser tweezers to produce topological defects with prescribed topological charge, such as pairs of a Saturn ring and an anti-ring, hyperbolic and radial hedgehogs on a fiber, as well as zero-charge loops. We study the entanglement and topological charge interaction between the topological defects of the fiber and sphere and we observe a huge variety of different entanglement topologies and defect-mediated elastic bindings. We explain all observed phenomena with simple topological rule: like topological charges repel each other and opposite topological charges attract. These binding mechanisms not only demonstrate the fascinating topology of nematic colloids, but also open a novel route to the assembly of very complex topological networks of fibers, spheres and other objects for applications in liquid crystal photonics.
DOI
Cell mechanics in biomedical cavitation
Qianxi Wang, Kawa Manmi, Kuo-Kang Liu
Studies on the deformation behaviours of cellular entities, such as coated microbubbles and liposomes subject to a cavitation flow, become increasingly important for the advancement of ultrasonic imaging and drug delivery. Numerical simulations for bubble dynamics of ultrasound contrast agents based on the boundary integral method are presented in this work. The effects of the encapsulating shell are estimated by adapting Hoff's model used for thin-shell contrast agents. The viscosity effects are estimated by including the normal viscous stress in the boundary condition. In parallel, mechanical models of cell membranes and liposomes as well as state-of-the-art techniques for quantitative measurement of viscoelasticity for a single cell or coated microbubbles are reviewed. The future developments regarding modelling and measurement of the material properties of the cellular entities for cutting-edge biomedical applications are also discussed.
DOI
Studies on the deformation behaviours of cellular entities, such as coated microbubbles and liposomes subject to a cavitation flow, become increasingly important for the advancement of ultrasonic imaging and drug delivery. Numerical simulations for bubble dynamics of ultrasound contrast agents based on the boundary integral method are presented in this work. The effects of the encapsulating shell are estimated by adapting Hoff's model used for thin-shell contrast agents. The viscosity effects are estimated by including the normal viscous stress in the boundary condition. In parallel, mechanical models of cell membranes and liposomes as well as state-of-the-art techniques for quantitative measurement of viscoelasticity for a single cell or coated microbubbles are reviewed. The future developments regarding modelling and measurement of the material properties of the cellular entities for cutting-edge biomedical applications are also discussed.
DOI
Micro- and nanotechnology for cell biophysics
Péter Galajda, Lóránd Kelemen, Gergely A. Végh
Procedures and methodologies used in cell biophysics have been improved tremendously with the revolutionary advances witnessed in the micro- and nanotechnology in the last two decades. With the advent of microfluidics it became possible to reduce laboratory-sized equipment to the scale of a microscope slide allowing massive parallelization of measurements with extremely low sample volume at the cellular level. Optical micromanipulation has been used to measure forces or distances or to alter the behavior of biological systems from the level of DNA to organelles or entire organisms. Among the main advantages is its non-invasiveness, giving researchers an invisible micro-hand to “touch” or “feel” the system under study, its freely and very often quickly adjustable experimental parameters such as wavelength, optical power or intensity distribution. Atomic force microscopy (AFM) opened avenues for in vitro biological applications concerning with single molecule imaging, cellular mechanics or morphology. As it can operate in liquid environment and at human body temperature, it became the most reliable and accurate nanoforce-tool in the research of cell biophysics. In this paper we review how the above three techniques help increase our knowledge in biophysics at the cellular level.
DOI
Procedures and methodologies used in cell biophysics have been improved tremendously with the revolutionary advances witnessed in the micro- and nanotechnology in the last two decades. With the advent of microfluidics it became possible to reduce laboratory-sized equipment to the scale of a microscope slide allowing massive parallelization of measurements with extremely low sample volume at the cellular level. Optical micromanipulation has been used to measure forces or distances or to alter the behavior of biological systems from the level of DNA to organelles or entire organisms. Among the main advantages is its non-invasiveness, giving researchers an invisible micro-hand to “touch” or “feel” the system under study, its freely and very often quickly adjustable experimental parameters such as wavelength, optical power or intensity distribution. Atomic force microscopy (AFM) opened avenues for in vitro biological applications concerning with single molecule imaging, cellular mechanics or morphology. As it can operate in liquid environment and at human body temperature, it became the most reliable and accurate nanoforce-tool in the research of cell biophysics. In this paper we review how the above three techniques help increase our knowledge in biophysics at the cellular level.
DOI
Plasmonic Coupling Dynamics of Silver Nanoparticles in an Optical Trap
Marc Blattmann and Alexander Rohrbach
We investigate binding and plasmonic coupling between optically trapped 80 nm silver spheres using a combination of spectroscopic sensing and 3D interferometric laser particle tracking on a 1 μs time scale. We demonstrate that nanoparticle coupling can be either spontaneous or induced by another particle through confinement of diffusion. We reveal ultrafast entries and exits of nanoparticles inside the optical trap, fast particle rearrangements before binding, and dimer formation allowing new insights into nanoparticle self-assembly.
DOI
We investigate binding and plasmonic coupling between optically trapped 80 nm silver spheres using a combination of spectroscopic sensing and 3D interferometric laser particle tracking on a 1 μs time scale. We demonstrate that nanoparticle coupling can be either spontaneous or induced by another particle through confinement of diffusion. We reveal ultrafast entries and exits of nanoparticles inside the optical trap, fast particle rearrangements before binding, and dimer formation allowing new insights into nanoparticle self-assembly.
DOI
Wednesday, December 2, 2015
‘Lissajous-like’ trajectories in optical tweezers
R. F. Hay, G. M. Gibson, S. H. Simpson, M. J. Padgett, and D. B. Phillips
When a microscopic particle moves through a low Reynolds number fluid, it creates a flow-field which exerts hydrodynamic forces on surrounding particles. In this work we study the ‘Lissajous-like’ trajectories of an optically trapped ‘probe’ microsphere as it is subjected to time-varying oscillatory hydrodynamic flow-fields created by a nearby moving particle (the ‘actuator’). We show a breaking of time-reversal symmetry in the motion of the probe when the driving motion of the actuator is itself time-reversal symmetric. This symmetry breaking results in a fluid-pumping effect, which arises due to the action of both a time-dependent hydrodynamic flow and a position-dependent optical restoring force, which together determine the trajectory of the probe particle. We study this situation experimentally, and show that the form of the trajectories observed is in good agreement with Stokesian dynamics simulations. Our results are related to the techniques of active micro-rheology and flow measurement, and also highlight how the mere presence of an optical trap can perturb the environment it is in place to measure.
DOI
When a microscopic particle moves through a low Reynolds number fluid, it creates a flow-field which exerts hydrodynamic forces on surrounding particles. In this work we study the ‘Lissajous-like’ trajectories of an optically trapped ‘probe’ microsphere as it is subjected to time-varying oscillatory hydrodynamic flow-fields created by a nearby moving particle (the ‘actuator’). We show a breaking of time-reversal symmetry in the motion of the probe when the driving motion of the actuator is itself time-reversal symmetric. This symmetry breaking results in a fluid-pumping effect, which arises due to the action of both a time-dependent hydrodynamic flow and a position-dependent optical restoring force, which together determine the trajectory of the probe particle. We study this situation experimentally, and show that the form of the trajectories observed is in good agreement with Stokesian dynamics simulations. Our results are related to the techniques of active micro-rheology and flow measurement, and also highlight how the mere presence of an optical trap can perturb the environment it is in place to measure.
DOI
Spin–orbit photonics
Filippo Cardano & Lorenzo Marrucci
Spin–orbit optical phenomena involve the interaction of the photon spin with the light wave propagation and spatial distribution, mediated by suitable optical media. Here we present a short overview of the emerging photonic applications that rely on such effects.
DOI
Spin–orbit optical phenomena involve the interaction of the photon spin with the light wave propagation and spatial distribution, mediated by suitable optical media. Here we present a short overview of the emerging photonic applications that rely on such effects.
DOI
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