Crossed fiber optic Bessel beams for curvilinear optofluidic transport of dielectric particles

Jongki Kim, Sungrae Lee, Yoonseob Jeong, Jun-Ki Kim, Yongmin Jung, Fabrice Merenda, Renè-Paul Salathè, Jeon-Soo Shin, and Kyunghwan Oh
Due to its unique non-diffracting and self-reconstructing nature, Bessel beams have been successfully adopted to trap multiple particles along the beam’s axial direction. However, prior bulk-optic based Bessel beams have a fundamental form-factor limitation for in situ, in-vitro, and in-vivo applications. Here we present a novel implementation of Fourier optics along a single strand of hybrid optical fiber in a monolithic manner that can generate pseudo Bessel beam arrays in two-dimensional space. We successfully demonstrate unique optofluidic transport of the trapped dielectric particles along a curvilinear optical route by multiplexing the fiber optic pseudo Bessel beams. The proposed technique can form a new building block to realize reconfigurable optofluidic transportation of particulates that can break the limitations of both prior bulk-optic Bessel beam generation techniques and conventional microfluidic channels.
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Optical trapping of NaYF4:Er3+,Yb3+ upconverting fluorescent nanoparticles

Patricia Haro, Blanca del Rosal, Laura Martínez Maestro, Rafik Naccache, John A. Capobianco, Kishan Dholakia, José García Solé, Emma Martín Rodríguez and Daniel Jaque García
We report on the first experimental observation of stable optical trapping of dielectric NaYF4:Er3+,Yb3+ upconverting fluorescent nanoparticles (~26 nm in diameter) using a continuous wave 980 nm single-beam laser. The laser serves both to optically trap and to excite visible luminescence from the nanoparticles. Sequential loading of individual nanoparticles into the trap is observed from the analysis of the emitted luminescence. We demonstrate that the trapping strength and the number of individual nanoparticles trapped is dictated by both the laser power and nanoparticle density. The possible contribution of thermal effects has been investigated by performing trapping experiments in heavy water in addition to distilled water. For the case of heavy water, thermal gradients are negligible and optical forces dominate the trap loading behaviour. The results provide a promising path towards real three dimensional manipulation of single NaYF4:Er3+,Yb3+ nanoparticles for precise fluorescence sensing in biophotonics experiments.
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Optical separation of droplets on a microfluidic platform

Jin Ho Jung, Kyung Heon Lee, Kang Soo Lee, Byung Hang Ha, Yong Suk Oh, Hyung Jin Sung
This paper describes the optical separation of microdroplets according to their refractive indices. The behavior of the droplets was characterized in terms of the optical force and the hydrodynamic effects present upon illumination of the droplets in a direction normal to the flow direction in a rectangular microfluidic channel. The optical forces acting on the droplets and the resultant droplet trajectories were analyzed and compared with the numerically predicted values. The relationship between the drag force and optical force was examined to understand the system performance properties in the context of screening applications involving the removal of unwanted droplets. Two species of droplets were compared for their photophoretic displacements by varying the illumination intensity. Because the optical forces exerted on the droplets were functions of the refractive indices and sizes of the droplets, a variety of chemical species could be separated simultaneously.
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Optical manipulation of Saccharomyces cerevisiae cells reveals that green light protection against UV irradiation is favored by low Ca2+ and requires intact UPR pathway

Ileana C. Farcasanu, Radu Mitrica, Ligia Cristache, Ioana Nicolau, Lavinia L. Ruta, Liliana Paslaru, Sorin Comorosan
Optical manipulation of Saccharomyces cerevisiae cells with high density green photons conferred protection against the deleterious effects of UV radiation. Combining chemical screening with UV irradiation of yeast cells, it was noted that the high density green photons relied on the presence of intact unfolded protein response (UPR) pathway to exert their protective effect and that the low Ca2+ conditions boosted the effect. UPR chemical inducers tunicamycin, dithiotreitol and calcium chelators augmented the green light effect in a synergic action against UV-induced damage. Photo-manipulation of cells was a critical factor since the maximum protection was achieved only when cells were pre-exposed to green light.
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Selective Adsorption of Functionalized Nanoparticles to Patterned Polymer Brush Surfaces and Its Probing with an Optical Trap

Annina Steinbach, Dr. Tobias Paust, Dr. Manuela Pluntke, Prof. Dr. Othmar Mart2, Prof. Dr. Dirk Volkmer
The site-specific attachment of nanoparticles is of interest for biomaterials or biosensor applications. Polymer brushes can be used to regulate this adsorption, so the conditions for selective adsorption of phosphonate-functionalized nanoparticles onto micropatterned polymer brushes with different functional groups are optimized. By choosing the strong polyelectrolytes poly(3-sulfopropyl methacrylate), poly(sulfobetaine methacrylate), and poly[2-(methacryloyloxy)ethyl trimethylammonium chloride], it is possible to direct the adsorption of nanoparticles to specific regions of the patterned substrates. A pH-dependent adsorption can be achieved by using the polycarboxylate brush poly(methacrylic acid) (PMAA) as substrate coating. On PMAA brushes, the nanoparticles switch from attachment to the brush regions to attachment to the grooves of a patterned substrate on changing the pH from 3 to 7. In this manner, patterned substrates are realized that assemble nanoparticles in pattern grooves, in polymer brush areas, or substrates that resist the deposition of the nanoparticles. The nanoparticle deposition can be directed in a pH-dependent manner on a weak polyelectrolyte, or is solely charge-dependent on strong polyelectrolytes. These results are correlated with surface potential measurements and show that an optical trap is a versatile method to directly probe interactions between nanoparticles and polymer brushes. A model for these interactions is proposed based on the optical trap measurements.

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High-Resolution Optical Tweezers for Single-Molecule Manipulation

Xinming Zhang, Lu Ma, and Yongli Zhang
Forces hold everything together and determine its structure and dynamics. In particular, tiny forces of 1-100 piconewtons govern the structures and dynamics of biomacromolecules. These forces enable folding, assembly, conformational fluctuations, or directional movements of biomacromolecules over sub-nanometer to micron distances. Optical tweezers have become a revolutionary tool to probe the forces, structures, and dynamics associated with biomacromolecules at a single-molecule level with unprecedented resolution. In this review, we introduce the basic principles of optical tweezers and their latest applications in studies of protein folding and molecular motors. We describe the folding dynamics of two strong coiled coil proteins, the GCN4-derived protein pIL and the SNARE complex. Both complexes show multiple folding intermediates and pathways. ATP-dependent chromatin remodeling complexes translocate DNA to remodel chromatin structures. The detailed DNA translocation properties of such molecular motors have recently been characterized by optical tweezers, which are reviewed here. Finally, several future developments and applications of optical tweezers are discussed. These past and future applications demonstrate the unique advantages of high-resolution optical tweezers in quantitatively characterizing complex multi-scale dynamics of biomacromolecules.

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Cluster formation in ferrofluids induced by holographic optical tweezers

Jan Masajada, Marcin Bacia, and Sławomir Drobczyński
Holographic optical tweezers were used to show the interaction between a strongly focused laser beam and magnetic nanoparticles in ferrofluid. When the light intensity was high enough, magnetic nanoparticles were removed from the beam center and formed a dark ring. The same behavior was observed when focusing vortex or Bessel beams. The interactions between two or more separated rings of magnetic nanoparticles created by independent optical traps were also observed.

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Interrogating Biology with Force: Single Molecule High-Resolution Measurements with Optical Tweezers

Marco Capitanio, Francesco S. Pavone
Single molecule force spectroscopy methods, such as optical and magnetic tweezers and atomic force microscopy, have opened up the possibility to study biological processes regulated by force, dynamics of structural conformations of proteins and nucleic acids, and load-dependent kinetics of molecular interactions. Among the various tools available today, optical tweezers have recently seen great progress in terms of spatial resolution, which now allows the measurement of atomic-scale conformational changes, and temporal resolution, which has reached the limit of the microsecond-scale relaxation times of biological molecules bound to a force probe. Here, we review different strategies and experimental configurations recently developed to apply and measure force using optical tweezers. We present the latest progress that has pushed optical tweezers’ spatial and temporal resolution down to today’s values, discussing the experimental variables and constraints that are influencing measurement resolution and how these can be optimized depending on the biological molecule under study.
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Laser Heating Tunability by Off-Resonant Irradiation of Gold Nanoparticles

Silvia Hormeño, Paula Gregorio-Godoy, Jorge Pérez-Juste, Luis M. Liz-Marzán, Beatriz H. Juárez, J. Ricardo Arias-Gonzalez
Temperature changes in the vicinity of a single absorptive nanostructure caused by local heating have strong implications in technologies such as integrated electronics or biomedicine. Herein, the temperature changes in the vicinity of a single optically trapped spherical Au nanoparticle encapsulated in a thermo-responsive poly(N-isopropylacrylamide) shell (Au@pNIPAM) are studied in detail. Individual beads are trapped in a counter-propagating optical tweezers setup at various laser powers, which allows the overall particle size to be tuned through the phase transition of the thermo-responsive shell. The experimentally obtained sizes measured at different irradiation powers are compared with average size values obtained by dynamic light scattering (DLS) from an ensemble of beads at different temperatures. The size range and the tendency to shrink upon increasing the laser power in the optical trap or by increasing the temperature for DLS agree with reasonable accuracy for both approaches. Discrepancies are evaluated by means of simple models accounting for variations in the thermal conductivity of the polymer, the viscosity of the aqueous solution and the absorption cross section of the coated Au nanoparticle. These results show that these parameters must be taken into account when considering local laser heating experiments in aqueous solution at the nanoscale. Analysis of the stability of the Au@pNIPAM particles in the trap is also theoretically carried out for different particle sizes.
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Microfluidic growth chambers with optical tweezers for full spatial single-cell control and analysis of evolving microbes

Christopher Probst, Alexander Grünberger, Wolfgang Wiechert, Dietrich Kohlheyer
Single-cell analysis in microfluidic systems has opened up new possibilities in biotechnological research enabling us to deal with large eukaryotic cells and even small bacteria. In particular, transient investigations in laminar flow or diffusive environments can be performed to unravel single cell behaviour. Up to now, most systems have been limited with respect to precise cell inoculation and sampling methods. Individual cell selection and manipulations have now been made possible by combining laser tweezers with microfluidic cell cultivation environments specifically tailored for micrometre-sized bacteria. Single cells were optically seeded into various micrometre-sized growth sites arranged in parallel. During cultivation, single-cell elongation, morphology and growth rates were derived from single cells and microcolonies of up to 500 cells. Growth of irradiated bacteria was not impaired by minimizing the exposed laser dosage as confirmed by exceptional growth rates. In fact, Escherichia coli exhibited doubling times of less than 20 min. For the first time, a filamentous Escherichia coli WT (MG1655) was safely relocated from its growing microcolony by laser manipulations. The cell was transferred to an empty cultivation spot allowing single-cell growth and morphology investigations. Contrary to previous discussions, the filamentous E. coli exhibited normal cell morphology and division after a few generations. This combination of optical tweezers and single-cell analysis in microfluidics adds a new degree of freedom to microbial single-cell analysis.
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Optical collection of multiple spheres in single tightly focused beams

Yongyin Cao, Lixue Chen, Weiqiang Ding, Fangkui Sun, Tongtong Zhu
The algorithm of multiple scattering is presented in this paper. The model of dynamic simulation and collision detection of multiple spheres is developed. The optical collection of multiple spheres in single tightly focused beams is simulated dynamically, considering the effects of Brownian motion of the spheres and the optical forces. The number of spheres with different parameters collected in a Gaussian beam is obtained using a program based on the T-matrix method and multiple scattering. Some interesting effects of the refractive index and size of spheres are revealed. The optical collection is explained physically in terms of optical binding. The method for modeling the collection of microspheres can be applied to the optical trapping and manipulation of multiple particles in general.
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Kinetics of Germination of Individual Spores of Geobacillus stearothermophilus as Measured by Raman Spectroscopy and Differential Interference Contrast Microscopy

Tingting Zhou, Zhiyang Dong, Peter Setlow, Yong-qing Li

Geobacillus stearothermophilus is a gram-positive, thermophilic bacterium, spores of which are very heat resistant. Raman spectroscopy and differential interference contrast microscopy were used to monitor the kinetics of germination of individual spores of G. stearothermophilus at different temperatures, and major conclusions from this work were as follows. 1) The CaDPA level of individual G. stearothermophilus spores was similar to that of Bacillus spores. However, the Raman spectra of protein amide bands suggested there are differences in protein structure in spores of G. stearothermophilus and Bacillus species. 2) During nutrient germination of G. stearothermophilus spores, CaDPA was released beginning after a lag time (Tlag) between addition of nutrient germinants and initiation of CaDPA release. CaDPA release was complete at Trelease, and ΔTrelease (Trelease – Tlag) was 1–2 min. 3) Activation by heat or sodium nitrite was essential for efficient nutrient germination of G. stearothermophilus spores, primarily by decreasing Tlag values. 4) Values of Tlag and Trelease were heterogeneous among individual spores, but ΔTrelease values were relatively constant. 5) Temperature had major effects on nutrient germination of G. stearothermophilus spores, as at temperatures below 65°C, average Tlag values increased significantly. 6) G. stearothermophilus spore germination with exogenous CaDPA or dodecylamine was fastest at 65°C, with longer Tlag values at lower temperatures. 7) Decoating of G. stearothermophilus spores slowed nutrient germination slightly and CaDPA germination significantly, but increased dodecylamine germination markedly. These results indicate that the dynamics and heterogeneity of the germination of individual G. stearothermophilus spores are generally similar to that of Bacillus species.
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Why Single-Beam Optical Tweezers Trap Gold Nanowires in Three Dimensions

Zijie Yan, Matthew Pelton, Leonid Vigderman, Eugene R. Zubarev, and Norbert F. Scherer
Understanding whether noble-metal nanostructures can be trapped optically and under what conditions will enable a range of applications that exploit their plasmonic properties. However, there are several nontrivial issues that first need to be resolved. A major one is that metal particles experience strong radiation pressure in optical beams, while stable optical trapping requires an attractive force greater than this radiation pressure. Therefore, it has generally been considered impossible to obtain sufficiently strong gradient forces using single-beam optical tweezers to trap relatively large metal nanostructures in three dimensions. Here we demonstrate that a single, tightly focused laser beam with a wavelength of 800 nm can achieve three-dimensional optical trapping of individual gold (Au) nanowires with lengths over 2 μm. Nanowires can be trapped by the beam at one of their ends, in which case they undergo significant angular fluctuations due to Brownian motion of the untrapped end. They can also be trapped close to their midpoints, in which case they are oriented approximately perpendicular to the light polarization direction. The behavior is markedly different from that of Ag nanowires with similar length and diameter, which cannot be trapped in three dimensions by a single focused Gaussian beam. Our results, including electrodynamics simulations that help to explain our experimental findings, suggest that the conventional wisdom, which holds that larger metal particles cannot be trapped, needs to be replaced with an understanding based on the details of plasmon resonances in the particles.
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RecG and UvsW catalyse robust DNA rewinding critical for stalled DNA replication fork rescue

Maria Manosas, Senthil K. Perumal, Piero Bianco, Felix Ritort, Stephen J. Benkovic & Vincent Croquette

Helicases that both unwind and rewind DNA have central roles in DNA repair and genetic recombination. In contrast to unwinding, DNA rewinding by helicases has proved difficult to characterize biochemically because of its thermodynamically downhill nature. Here we use single-molecule assays to mechanically destabilize a DNA molecule and follow, in real time, unwinding and rewinding by two DNA repair helicases, bacteriophage T4 UvsW and Escherichia coli RecG. We find that both enzymes are robust rewinding enzymes, which can work against opposing forces as large as 35 pN, revealing their active character. The generation of work during the rewinding reaction allows them to couple rewinding to DNA unwinding and/or protein displacement reactions central to the rescue of stalled DNA replication forks. The overall results support a general mechanism for monomeric rewinding enzymes.

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Microrheological Investigations in Ionic Liquids Using Optical Trapping Techniques

Richard D. Dear , Emma K. Worrall , William D. Gault , and Grant A. D. Ritchie
In this paper, we demonstrate optical trapping of melamine particles (d ≈ 2.3 μm) within the pure ionic liquid ethylammonium nitrate (EAN) and show the first microrheological investigations of these important compounds using this technique. By analyzing the power spectrum of a particle trapped in EAN, we monitor the variation in viscous drag that it experiences in proximity to the sample coverslip, showing excellent agreement with Faxén’s law. We also demonstrate hydrodynamic coupling between pairs of trapped particles. Finally, we explore temperature-dependent viscosity changes in μL samples of EAN as a further example of microrheological investigations of ILs.

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Single-Molecule Folding Mechanism of an EF-Hand Neuronal Calcium Sensor

Pétur O. Heidarsson, Mariela R. Otazo, Luca Bellucci, Alessandro Mossa, Alberto Imparato, Emanuele Paci, Stefano Corni, Rosa Di Felice, Birthe B. Kragelund, Ciro Cecconi
EF-hand calcium sensors respond structurally to changes in intracellular Ca2+ concentration, triggering diverse cellular responses and resulting in broad interactomes. Despite impressive advances in decoding their structure-function relationships, the folding mechanism of neuronal calcium sensors is still elusive. We used single-molecule optical tweezers to study the folding mechanism of the human neuronal calcium sensor 1 (NCS1). Two intermediate structures induced by Ca2+ binding to the EF-hands were observed during refolding. The complete folding of the C domain is obligatory for the folding of the N domain, showing striking interdomain dependence. Molecular dynamics results reveal the atomistic details of the unfolding process and rationalize the different domain stabilities during mechanical unfolding. Through constant-force experiments and hidden Markov model analysis, the free energy landscape of the protein was reconstructed. Our results emphasize that NCS1 has evolved a remarkable complex interdomain cooperativity and a fundamentally different folding mechanism compared to structurally related proteins.
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Spectral tuning of lasing emission from optofluidic droplet microlasers using optical stretching

Mehdi Aas, Alexandr Jonáš, Alper Kiraz, Oto Brzobohatý, Jan Ježek, Zdeněk Pilát, and Pavel Zemánek
We introduce tunable optofluidic microlasers based on active optical resonant cavities formed by optically stretched, dye-doped emulsion droplets confined in a dual-beam optical trap. To achieve tunable dye lasing, optically pumped droplets of oil dispersed in water are stretched by light in the dual-beam trap. Subsequently, resonant path lengths of whispering gallery modes (WGMs) propagating in the droplet are modified, leading to shifts in the microlaser emission wavelengths. Using this technique, we present all-optical, almost reversible spectral tuning of the lasing WGMs and show that the direction of tuning depends on the position of the pump beam focus on the droplet. In addition, we study the effects of temperature changes on the spectral position of lasing WGMs and demonstrate that droplet heating leads to red-tuning of the droplet lasing wavelength.
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Independent and simultaneous three-dimensional optical trapping and imaging

Maya Yevnin, Dror Kasimov, Yael Gluckman, Yuval Ebenstein, and Yael Roichman

Combining imaging and control of multiple micron-scaled objects in three dimensions opens up new experimental possibilities such as the fabrication of colloidal-based photonic devices, as well as high-throughput studies of single cell dynamics. Here we utilize the dual-objectives approach to combine 3D holographic optical tweezers with a spinning-disk confocal microscope. Our setup is capable of trapping multiple different objects in three dimensions with lateral and axial accuracy of 8 nm and 20 nm, and precision of 20 nm and 200 nm respectively, while imaging them in four different fluorescence channels. We demonstrate fabrication of ordered two-component and three dimensional colloidal arrays, as well as trapping of yeast cell arrays. We study the kinetics of the division of yeast cells within optical traps, and find that the timescale for division is not affected by trapping.

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Optical trapping of fluorescent beads

Jitendra Bhatt, Ashok Kumar, R. P. Singh and S. N.A. Jaaffrey

Optical tweezers have been successfully used to trap a variety of particles and biological specimens for numerous applications. Particles which are reflective as well as absorbing could be trapped using beams such as optical vortex. Here we give the details of our efforts to trap fluorescent microparticles. We have set up an optical trap for these fluorescent microparticles using holographic optical tweezers; we observe that it is not possible to trap fluorescent microparticles with a Gaussian laser beam or a hollow beam. However, as the fluorescence of these particles gets degraded they could be trapped in custom-made holographic tweezers. Moreover, when a fluorescent particle is brought in the trap containing stably trapped non-fluorescent particle, the stably trapped non-fluorescent particle also escapes from the trap.
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Electrochemical Detection of Single Microbeads Manipulated by Optical Tweezers in the Vicinity of Ultramicroelectrodes

Emmanuel Suraniti, Frédéric Kanoufi, Charlie Gosse, Xuan Zhao, Rumiana Dimova, Bernard Pouligny, and Neso Sojic
Latex micrometric beads are manipulated by optical tweezers in the vicinity of an ultramicroelectrode (UME). They are optically trapped in solution and approached the electrode surface. After the electrochemical measurement, they are optically removed from the surface. The residence time of the particle on the electrode is thus controlled by the optical tweezers. The detection is based on diffusional hindrance by the insulating objects which alters the fluxes of the redox Ru(NH3)63+ species toward the UME and thus its mass-transfer limited current. We have optically deposited successively 1, 2, and 3 beads of 3-μm radius on the UME surface, and we have recorded the variations of the current depending on their landing locations that were optically controlled. Finally we decreased the current by partially blocking the electroactive surface with a six-bead assembly. The variation of the steady-state current and the approach curves allow for the indirect electrochemical localization of the bead in the vicinity of the UME, not only when the bead is in contact but also when it is levitated at distances lower than the UME radius. These experiments show that single particles or more complex structures may be manipulated in situ in a contactless mode near the UME surface. From comparison with simulations, the electrochemical detection affords an indirect localization of the object in the UME environment. The developed approach offers a potential application for interrogating the electrochemical activity of single cells and nanoparticles.
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Single-Molecule Unzipping Force Analysis of HU–DNA Complexes

Dr. Remus T. Dame, Dr. Michael A. Hall, Dr. Michelle D. Wang
The genome of bacteria is organized and compacted by the action of nucleoid-associated proteins. These proteins are often present in tens of thousands of copies and bind with low specificity along the genome. DNA-bound proteins thus potentially act as roadblocks to the progression of machinery that moves along the DNA. In this study, we have investigated the effect of histone-like protein from strain U93 (HU), one of the key proteins involved in shaping the bacterial nucleoid, on DNA helix stability by mechanically unzipping single dsDNA molecules. Our study demonstrates that individually bound HU proteins have no observable effect on DNA helix stability, whereas HU proteins bound side-by-side within filaments increase DNA helix stability. As the stabilizing effect is small compared to the power of DNA-based motor enzymes, our results suggest that HU alone does not provide substantial hindrance to the motor's progression in vivo.

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Surface phonon-polariton enhanced optical forces in silicon carbide nanostructures

Dongfang Li, Nabil M. Lawandy, and Rashid Zia
The enhanced optical forces induced by surface phonon-polariton (SPhP) modes are investigated in different silicon carbide (SiC) nanostructures. Specifically, we calculate optical forces using the Maxwell stress tensor for three different geometries: spherical particles, slab waveguides, and rectangular waveguides. We show that SPhP modes in SiC can produce very large forces, more than one order of magnitude larger than the surface plasmon-polariton (SPP) forces in analogous metal nanostructures. The material and geometric basis for these large optical forces are examined in terms of dispersive permittivity, separation distance, and operating wavelength.
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Centrosome repositioning in T cells is biphasic and driven by microtubule end-on capture-shrinkage

Jason Yi, Xufeng Wu, Andrew H. Chung, James K. Chen, Tarun M. Kapoor, and John A. Hammer
T cells rapidly reposition their centrosome to the center of the immunological synapse (IS) to drive polarized secretion in the direction of the bound target cell. Using an optical trap for spatial and temporal control over target presentation, we show that centrosome repositioning in Jurkat T cells exhibited kinetically distinct polarization and docking phases and required calcium flux and signaling through both the T cell receptor and integrin to be robust. In “frustrated” conjugates where the centrosome is stuck behind the nucleus, the center of the IS invaginated dramatically to approach the centrosome. Consistently, imaging of microtubules during normal repositioning revealed a microtubule end-on capture-shrinkage mechanism operating at the center of the IS. In agreement with this mechanism, centrosome repositioning was impaired by inhibiting microtubule depolymerization or dynein. We conclude that dynein drives centrosome repositioning in T cells via microtubule end-on capture-shrinkage operating at the center of the IS and not cortical sliding at the IS periphery, as previously thought.
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Probing cell biophysical behavior based on actin cytoskeleton modeling and stretching manipulation with optical tweezers

Kaiqun Wang, Jinping Cheng, Shuk Han Cheng, and Dong Sun
This letter presents an approach to utilizing the actin cytoskeleton model and optical tweezers technology to probe the distinct underlying F-actin remodeling mechanism and showing quantitatively how cell mechanical behavior is associated with alterations in the cell functions. The structural parameters of F-actin were extracted by fitting the modeling results with the experimental results obtained by cell stretching manipulation. Alterations of cell mechanical behaviors under distinct diseased cellular stages were further interpreted. Jurkat and K562 cells were used as sample cells. This letter successfully illustrates the correlation of the cell mechanical behavior and cell functional alterations in a quantitative way.

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Maximizing information content of single-molecule FRET experiments: multi-color FRET and FRET combined with force or torque

Sungchul Hohng, Sanghwa Lee, Jinwoo Lee and Myung Hyun Jo
Since its first demonstration about twenty years ago, single-molecule fluorescence resonance energy transfer (FRET) has undergone remarkable technical advances. In this tutorial review, we will discuss two technical advances that increase the information content of the single-molecule FRET measurements: single-molecule multi-color FRET and single-molecule FRET combined with force or torque. Our expectations for future developments will be briefly discussed at the end.
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Mechanochemical Properties of Individual Human Telomeric RNA (TERRA) G-Quadruplexes

Philip M. Yangyuoru, Dr. Amy Y. Q. Zhang, Zhe Shi, Deepak Koirala, Prof. Shankar Balasubramanian, Dr. Hanbin Mao

Potential functions: By following the unfolding and refolding of individual human RNA telomeric (TERRA) G-quadruplexes (GQs) in laser tweezers, the mechanical stability and transition kinetics of RNA GQs are obtained. Comparison between TERRA and DNA GQs suggests their different regulatory capacities for processes associated with human telomeres.

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Shaping of light beams along curves in three dimensions

José A. Rodrigo, Tatiana Alieva, Eugeny Abramochkin, and Izan CastroWe present a method for efficient and versatile generation of beams whose intensity and phase are prescribed along arbitrary 3D curves. It comprises a non-iterative beam shaping technique that does not require solving inversion problems of light propagation. The generated beams have diffraction-limited focusing with high intensity and controlled phase gradients useful for applications such as laser micro-machining and optical trapping. Its performance and feasibility are experimentally demonstrated on several examples including multiple trapping of micron-sized particles.
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Automated Manipulation of Biological Cells Using Gripper Formations Controlled By Optical Tweezers

Sagar Chowdhury, Atul Thakur, Petr Švec, Chenlu Wang, Wolfgang Losert, and Satyandra K. Gupta
The capability of noninvasive and precise micromanipulation of sensitive, living cells is necessary for understanding their underlying biological processes. Optical tweezers (OT) is an effective tool that uses highly focused laser beams for accurate manipulation of cells and dielectric beads at microscale. However, direct exposure of the laser beams on the cells can negatively influence their behavior or even cause a photo-damage. In this paper, we introduce a control and planning approach for automated, indirect manipulation of cells using silica beads arranged into gripper formations. The developed approach employs path planning and feedback control for efficient, collision-free transport of a cell between two specified locations. The planning component of the approach computes a path that explicitly respects the nonholonomic constraints of the gripper formations. The feedback control component ensures stable tracking of the path by manipulating the cell using a set of predefined maneuvers. We demonstrate the effectiveness of the approach by transporting a yeast cell using four different types of gripper formations along collision-free paths on our OT setup. We analyzed the performance of the proposed gripper formations with respect to their maximum transport speeds and the laser intensity experienced by the cell that depends on the laser power used.
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PICH: A DNA Translocase Specially Adapted for Processing Anaphase Bridge DNA

Andreas Biebricher, Seiki Hirano, Jacqueline H. Enzlin, Nicola Wiechens, Werner W. Streicher, Diana Huttner, Lily H.-C. Wang, Erich A. Nigg, Tom Owen-Hughes, Ying Liu, Erwin Peterman, Gijs J.L. Wuite, Ian D. HicksonThe Plk1-interacting checkpoint helicase (PICH) protein localizes to ultrafine anaphase bridges (UFBs) in mitosis alongside a complex of DNA repair proteins, including the Bloom’s syndrome protein (BLM). However, very little is known about the function of PICH or how it is recruited to UFBs. Using a combination of microfluidics, fluorescence microscopy, and optical tweezers, we have defined the properties of PICH in an in vitro model of an anaphase bridge. We show that PICH binds with a remarkably high affinity to duplex DNA, resulting in ATP-dependent protein translocation and extension of the DNA. Most strikingly, the affinity of PICH for binding DNA increases with tension-induced DNA stretching, which mimics the effect of the mitotic spindle on a UFB. PICH binding also appears to diminish force-induced DNA melting. We propose a model in which PICH recognizes and stabilizes DNA under tension during anaphase, thereby facilitating the resolution of entangled sister chromatids.

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Cavity-enhanced optical trapping of bacteria using a silicon photonic crystal

Thijs van Leest and Jacob Caro

On-chip optical trapping and manipulation of cells based on the evanescent field of photonic structures is emerging as a promising technique, both in research and for applications in broader context. Relying on mass fabrication techniques, the involved integration of photonics and microfluidics allows control of both the flow of light and water on the scale of interest in single cell microbiology. In this paper, we demonstrate for the first time optical trapping of single bacteria (B. subtilis and E. coli) using photonic crystal cavities for local enhancement of the evanescent field, as opposed to the synthetic particles used so far. Three types of cavities (H0, H1 and L3) are studied, embedded in a planar photonic crystal and optimized for coupling to two collinear photonic crystal waveguides. The photonic crystals are fabricated on a silicon-on-insulator chip, onto which a fluidic channel is created as well. For each of the cavities we clearly demonstrate optical trapping of bacteria when they are pumped at the resonance frequency (around 1550 nm), in spite of their low index contrast w.r.t. water. By tracking the confined Brownian motion of B. subtilis spores in the traps using recorded microscope observations, we derive strong in-plane trap stiffnesses of about 7.6 pN/nm/W. The values found agree very well with calculations based on the Maxwell stress tensor for the force and finite-difference time-domain simulations of the fields for the fabricated cavity geometries. We envision that our lab-on-a-chip with photonic crystal traps opens up new application directions, e.g. immobilization of single bio-objects such as mammalian cells and bacteria under controlled conditions for optical microscopy studies.
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Non-spherical particles for optical trap assisted nanopatterning

Y-C Tsai, R Fardel, M M Panczyk, E M Furst and C B Arnold
Optical trap assisted nanopatterning is a laser direct-write technique that uses an optically trapped microsphere as a near-field objective. The type of feature that one can create with this technique depends on several factors, one of which is the shape of the microbead. In this paper, we examine how the geometry of the bead affects the focus of the light through a combination of experiments and simulations. We realize nanopatterning using non-spherical dielectric particles to shape the light–material interaction. We model the resulting nanoscale features with a finite difference time domain simulation and obtain very good agreement with the experiments. This work opens the way to systematic engineering of the microparticle geometry in order to tailor the near-field focus to specific nanopatterning applications.
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Pico-Force Optical Exchange (pico-FOX): Utilizing Optical Forces Applied to an Orthogonal Electroosmotic Flow for Particulate Enrichment from Mixed Sample Streams

Sarah J. R. Staton, Soo Y. Kim, Sean J. Hart, Greg E. Collins, and Alex Terray
Results are reported from a combined optical force and electrokinetic microfluidic device that separates individual particulates from molecular components in a mixed sample stream. A pico-Newton optical force was applied to an orthogonal electroosmotic flow carrying a hydrodynamically pinched, mixed sample, resulting in the separation of the various particles from the sample stream. Different combinations of polystyrene, PMMA, and silica particles with a commercially available dye were utilized to test the different separation modes available, from purely optical force to combined optical and electrophoretic forces. The impact of various particle properties on particle separation and separation efficiency were explored, including size (2, 6, 10 μm), refractive index, and electrophoretic mobility. Particle addressability was achieved by moving particles to different outlets on the basis of particle size, refractive index, and electrophoretic differences. Separations of 6 and 10 μm polystyrene particles led to only 3% particle contamination in the original sample stream and interparticle type enrichment levels >80%. The unique addressability of three different particle materials (polystyrene, PMMA, and silica) of the same size (2 μm) led to each being separated into a unique outlet without measurable contamination of the other particle types using optical force and electrophoretic mobility. In addition to particle separation, the device was able to minimize dye diffusion, leading to >95% dye recovery. This combined platform would have applications for noninvasive sample preparation of mixed molecular/particulate systems for mating with traditional analytics as well as efficient removal of harmful, degrading components from complex mixtures.
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“Red Tweezers:” Fast, customisable hologram generation for optical tweezers

Richard W. Bowman, Graham M. Gibson, Anna Linnenberger, David B. Phillips, James A. Grieve, David M. Carberry, Steven Serati, Mervyn J. Miles, Miles J. Padgett

Holographic Optical Tweezers (HOT) are a versatile way of manipulating microscopic particles in 3D. However, their ease of use has been hampered by the computational load of calculating the holograms, resulting in an unresponsive system. We present a program for generating these holograms on a consumer Graphics Processing Unit (GPU), coupled to an easy-to-use interface in LabVIEW (National Instruments). This enables a HOT system to be set up without writing any additional code, as well as providing a platform enabling the fast generation of other holograms. The GPU engine calculates holograms over 300 times faster than the same algorithm running on a quad core CPU. The hologram algorithm can be altered on-the-fly without recompiling the program, allowing it to be used to control Spatial Light Modulators in any situation where the hologram can be calculated in a single pass. The interface has also been rewritten to take advantage of new features in LabVIEW 2010. It is designed to be easily modified and extended to integrate with hardware other than our own.
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The growth speed of microtubules with XMAP215-coated beads coupled to their ends is increased by tensile force

Anastasiya Trushko, Erik Schäffer, and Jonathon Howard
The generation of pulling and pushing forces is one of the important functions of microtubules, which are dynamic and polarized structures. The ends of dynamic microtubules are able to form relatively stable links to cellular structures, so that when a microtubule grows it can exert a pushing force and when it shrinks it can exert a pulling force. Microtubule growth and shrinkage are tightly regulated by microtubule-associated proteins (MAPs) that bind to microtubule ends. Given their localization, MAPs may be exposed to compressive and tensile forces. The effect of such forces on MAP function, however, is poorly understood. Here we show that beads coated with the microtubule polymerizing protein XMAP215, the Xenopus homolog of Dis1 and chTOG, are able to link stably to the plus ends of microtubules, even when the ends are growing or shrinking; at growing ends, the beads increase the polymerization rate. Using optical tweezers, we found that tensile force further increased the microtubule polymerization rate. These results show that physical forces can regulate the activity of MAPs. Furthermore, our results show that XMAP215 can be used as a handle to sense and mechanically manipulate the dynamics of the microtubule tip.
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Dynein Interacts with the Neural Cell Adhesion Molecule (NCAM180) to Tether Dynamic Microtubules and Maintain Synaptic Density in Cortical Neurons

Eran Perlson, Adam G. Hendricks, Jacob E. Lazarus, Keren Ben-Yaakov, Tal Gradus, Mariko Tokito and Erika L. F. Holzbaur
Cytoplasmic dynein is well-characterized as an organelle motor, but dynein also acts to tether and stabilize dynamic microtubule plus-ends in vitro. Here we identify a novel and direct interaction between dynein and the 180 kDa isoform of the neural cell adhesion molecule NCAM. Optical trapping experiments indicate that dynein bound to beads via the NCAM180 interaction domain can tether projecting microtubule plus-ends. Live cell assays indicate that the NCAM180-dependent recruitment of dynein to the cortex leads to the selective stabilization of microtubules projecting to NCAM180 patches at the cell periphery. The dynein-NCAM180 interaction also enhances cell-cell adhesion in heterologous cell assays. Dynein and NCAM180 co-precipitate from mouse brain extract and from synaptosomal fractions, consistent with an endogenous interaction in neurons. Thus, we examined microtubule dynamics and synaptic density in primary cortical neurons. We find that depletion of NCAM, inhibition of the dynein-NCAM180 interaction or dampening of microtubule dynamics with low dose nocodazole all result in significant decreases in synaptic density. Based on these observations, we propose a working model for the role of dynein at the synapse, in which the anchoring of the motor to the cortex via binding to an adhesion molecule mediates the tethering of dynamic microtubule plus-ends to potentiate synaptic stabilization.
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Kinetochore kinesin CENP-E is a processive bi-directional tracker of dynamic microtubule tips

Nikita Gudimchuk, Benjamin Vitre, Yumi Kim, Anatoly Kiyatkin, Don W. Cleveland, Fazly I. Ataullakhanov & Ekaterina L. Grishchuk
During vertebrate mitosis, the centromere-associated kinesin CENP-E (centromere protein E) transports misaligned chromosomes to the plus ends of spindle microtubules. Subsequently, the kinetochores that form at the centromeres establish stable associations with microtubule ends, which assemble and disassemble dynamically. Here we provide evidence that after chromosomes have congressed and bi-oriented, the CENP-E motor continues to play an active role at kinetochores, enhancing their links with dynamic microtubule ends. Using a combination of single-molecule approaches and laser trapping in vitro, we demonstrate that once reaching microtubule ends, CENP-E converts from a lateral transporter into a microtubule tip-tracker that maintains association with both assembling and disassembling microtubule tips. Computational modelling of this behaviour supports our proposal that CENP-E tip-tracks bi-directionally through a tethered motor mechanism, which relies on both the motor and tail domains of CENP-E. Our results provide a molecular framework for the contribution of CENP-E to the stability of attachments between kinetochores and dynamic microtubule ends.
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Femtosecond Pulse-Width Dependent Trapping and Directional Ejection Dynamics of Dielectric Nanoparticles

Wei-Yi Chiang, Anwar Usman, and Hiroshi Masuhara
We demonstrate that laser pulse duration, which determines its impulsive peak power, is an effective parameter to control the number of optically trapped dielectric nanoparticles, their ejections along the directions perpendicular to polarization vector, and their migration distances from the trapping site. This ability to controllably confine and eject the nanoparticle is explained by pulse width-dependent optical forces exerted on nanoparticles in the trapping site and ratio between the repulsive and attractive forces. We also show that the directional ejections occur only when the number of nanoparticles confined in the trapping site exceeds a definite threshold. We interpret our data by considering the formation of transient assembly of the optically confined nanoparticles, partial ejection of the assembly, and subsequent filling of the trapping site. The understanding of optical trapping and directional ejections by ultrashort laser pulses paves the way to optically controlled manipulation and sorting of nanoparticles.
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Enumerating virus-like particles in an optically concentrated suspension by fluorescence correlation spectroscopy

Yi Hu, Xuanhong Cheng, and H. Daniel Ou-Yang
Fluorescence correlation spectroscopy (FCS) is one of the most sensitive methods for enumerating low concentration nanoparticles in a suspension. However, biological nanoparticles such as viruses often exist at a concentration much lower than the FCS detection limit. While optically generated trapping potentials are shown to effectively enhance the concentration of nanoparticles, feasibility of FCS for enumerating field-enriched nanoparticles requires understanding of the nanoparticle behavior in the external field. This paper reports an experimental study that combines optical trapping and FCS to examine existing theoretical predictions of particle concentration. Colloidal suspensions of polystyrene (PS) nanospheres and HIV-1 virus-like particles are used as model systems. Optical trapping energies and statistical analysis are used to discuss the applicability of FCS for enumerating nanoparticles in a potential well produced by a force field.
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Heartbeat-Driven Pericardiac Fluid Forces Contribute to Epicardium Morphogenesis

Marina Peralta, Emily Steed, Sébastien Harlepp, Juan Manuel González-Rosa, Fabien Monduc, Ana Ariza-Cosano, Alfonso Cortés, Teresa Rayón, Jose-Luis Gómez-Skarmeta, Agustín Zapata, Julien Vermot, Nadia Mercader
Hydrodynamic forces play a central role in organ morphogenesis. The role of blood flow in shaping the developing heart is well established, but the role of fluid forces generated in the pericardial cavity surrounding the heart is unknown. Mesothelial cells lining the pericardium generate the proepicardium (PE), the precursor cell population of the epicardium, the outer layer covering the myocardium, which is essential for its maturation and the formation of the heart valves and coronary vasculature. However, there is no evidence from in vivo studies showing how epicardial precursor cells reach and attach to the heart.
Using optical tools for real-time analysis in the zebrafish, including high-speed imaging and optical tweezing, we show that the heartbeat generates pericardiac fluid advections that drive epicardium formation. These flow forces trigger PE formation and epicardial progenitor cell release and motion. The pericardial flow also influences the site of PE cell adhesion to the myocardium. We additionally identify novel mesothelial sources of epicardial precursors and show that precursor release and adhesion occur both through pericardiac fluid advections and through direct contact with the myocardium.
Two hydrodynamic forces couple cardiac development and function: first, blood flow inside the heart, and second, the pericardial fluid advections outside the heart identified here. This pericardiac fluid flow is essential for epicardium formation and represents the first example of hydrodynamic flow forces controlling organogenesis through an action on mesothelial cells.
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Shear strengthens fibrin: the knob-hole interactions display “catch-slip” kinetics

R. I. Litvinov, J. W. Weisel
The polymerization of fibrin occurs primarily via intermolecular non-covalent binding between knobs ‘A’ exposed by cleavage of fibrinopeptides A in the Aα chains of fibrinogen and holes ‘a’ constitutively present in the γ chains [1]. We have previously investigated the interactions between knobs ‘A’ and holes ‘a’ at the single-molecule level and have found that they are strong and stable [2] with complex forced unbinding mechanisms [3]. The structural complexity and thermodynamics of these interactions must be reflected in the non-trivial dynamics of the mechanical response of the knob-hole bond to stress. By applying an original model system to quantify the mechanical dissociation of individual knob-hole complexes, we revealed an unusual strengthening of A:a knob-hole bonds in response to increasing pulling force, referred to as a “catch” bond.

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Three-dimensional quasicrystalline photonic material with five-fold planar symmetry for visible and infrared wavelengths by holographic assembly of quasicrystalline photonic heterostructures

Zaven Ovanesyan, Pushpa Raj Pudasaini, Ajithkumar Gangadharan, and Marcelo Marucho

In this paper, we investigate three-dimensional (3D) band gap properties of quasiperiodic structure. We successfully demonstrate the fabrication of a 3D dielectric quasicrystalline heterostructures with five-fold planar symmetry using the holographic optical tweezers technique. Light transmitted through this quasicrystal is collected using the spatially resolved optical spectroscopy technique for both visible and infrared wavelength bandwidths in a far-field region. We investigate and analyze the transmission spectra for the same wavelength bandwidths in a near-field region by using computer simulations. The computational modeling indicates that for both TE and TM modes of propagating light in the XY plane there is a clear transmission band-gap of around 50 nm wide centered at 650 nm. This indicates that there is a rotational symmetry in the constructed quasicrystal along its XY plane. Future directions and applications are discussed.
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Torsionally constrained DNA for single-molecule assays: an efficient, ligation-free method

D. Hern Paik, Violet A. Roskens and Thomas T. Perkins

Controlled twisting of individual, double-stranded DNA molecules provides a unique method to investigate the enzymes that alter DNA topology. Such twisting requires a single DNA molecule to be torsionally constrained. This constraint is achieved by anchoring the opposite ends of the DNA to two separate surfaces via multiple bonds. The traditional protocol for making such DNA involves a three-way ligation followed by gel purification, a laborious process that often leads to low yield both in the amount of DNA and the fraction of molecules that is torsionally constrained. We developed a simple ligation-free procedure for making torsionally constrained DNA via polymerase chain reaction (PCR). This PCR protocol used two ‘megaprimers’, 400-base-pair long double-stranded DNA that were labelled with either biotin or digoxigenin. We obtained a relatively high yield of gel-purified DNA (∼500 ng/100 µl of PCR reaction). The final construct in this PCR-based method contains only one labelled strand in contrast to the traditional construct in which both strands of the DNA are labelled. Nonetheless, we achieved a high yield (84%) of torsionally constrained DNA when measured using an optical-trap-based DNA-overstretching assay. This protocol significantly simplifies the application and adoption of torsionally constrained assays to a wide range of single-molecule systems.

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Cavity cooling of an optically levitated submicron particle

Nikolai Kiesel, Florian Blaser, Uroš Delić, David Grass, Rainer Kaltenbaek, and Markus Aspelmeyer
The coupling of a levitated submicron particle and an optical cavity field promises access to a unique parameter regime both for macroscopic quantum experiments and for high-precision force sensing. We report a demonstration of such controlled interactions by cavity cooling the center-of-mass motion of an optically trapped submicron particle. This paves the way for a light–matter interface that can enable room-temperature quantum experiments with mesoscopic mechanical systems.
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ear-field multiple optical trapping using high order axially symmetric polarized beams

Zhehai Zhou, Qiaofeng Tan, Changxi Yang, Lianqing Zhu

The near-field multiple optical trapping using high order axially symmetric polarized beams (ASPBs) is studied for the first time. First, a near-field optical trapping scheme is proposed based on the Kretschmann–Raether configuration, and surface plasmon polaritons (SPPs) field distributions excited by incident ASPBs are calculated, which present a multi-focal-spot pattern and the size of spots is much smaller than that of the diffraction limitation. Then, the gradient forces on Rayleigh dielectric particles formed by the multi-focal-spot focused field are computed, which indicates that multiple ultra-small particles with the refractive index higher than that of the ambient medium can be trapped simultaneously on the metal surface. The number and size of trapped particles can be manipulated by flexibly modifying the polarization order of incident beams, which is expected to enhance the capability of traditional optical trapping systems and provide a solution for massively parallel optical trapping of nanometer-sized particles.

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STED nanoscopy combined with optical tweezers reveals protein dynamics on densely covered DNA

Iddo Heller, Gerrit Sitters, Onno D Broekmans, Géraldine Farge, Carolin Menges, Wolfgang Wende, Stefan W Hell, Erwin J G Peterman & Gijs J L Wuite
Dense coverage of DNA by proteins is a ubiquitous feature of cellular processes such as DNA organization, replication and repair. We present a single-molecule approach capable of visualizing individual DNA-binding proteins on densely covered DNA and in the presence of high protein concentrations. Our approach combines optical tweezers with multicolor confocal and stimulated emission depletion (STED) fluorescence microscopy. Proteins on DNA are visualized at a resolution of 50 nm, a sixfold resolution improvement over that of confocal microscopy. High temporal resolution (<50 ms) is ensured by fast one-dimensional beam scanning. Individual trajectories of proteins translocating on DNA can thus be distinguished and tracked with high precision. We demonstrate our multimodal approach by visualizing the assembly of dense nucleoprotein filaments with unprecedented spatial resolution in real time. Experimental access to the force-dependent kinetics and motility of DNA-associating proteins at biologically relevant protein densities is essential for linking idealized in vitro experiments with the in vivo situation.

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Optical Vortex Induced Rotation of Silver Nanowires

Zijie Yan and Norbert F. Scherer
Optical manipulation of metal nanowires offers the possibility to control the position, orientation, and associated motions of individual nanowires, particularly by utilizing their plasmonic properties. Here, we demonstrate that the orbital angular momentum of photons in Laguerre–Gauss (optical vortex) beams can induce rotation of single silver (Ag) nanowires with lengths of over 10 μm that are lying on (in molecular proximity to) a dielectric surface. We show that the rotation dynamics are governed by plasmonic interactions of the Ag nanowires with linearly polarized light, which yield a sinusoidal optical torque that causes angular acceleration. These results provide important information to understand the angular dependence of plasmonic nanowire–light interactions and extend the repertoire to realize applications in plasmonic lab-on-a-chip systems.
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Nanolithography by Plasmonic Heating and Optical Manipulation of Gold Nanoparticles

Michael Fedoruk, Marco Meixner, Sol Carretero-Palacios, Theobald Lohmüller, and Jochen Feldmann

Noble-metal particles feature intriguing optical properties, which can be utilized to manipulate them by means of light. Light absorbed by gold nanoparticles, for example, is very efficiently converted into heat, and single particles can thus be used as a fine tool to apply heat to a nanoscopic area. At the same time, gold nanoparticles are subject to optical forces when they are irradiated with a focused laser beam, which renders it possible to print, manipulate, and optically trap them in two and three dimensions. Here, we demonstrate how these properties can be used to control the polymerization reaction and thermal curing of polydimethylsiloxane (PDMS) at the nanoscale and how these findings can be applied to synthesize polymer nanostructures such as particles and nanowires with subdiffraction limited resolution.
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Linear momentum increase and negative optical forces at dielectric interface

Veerachart Kajorndejnukul, Weiqiang Ding, Sergey Sukhov, Cheng-Wei Qiu & Aristide Dogariu
Light carries momenta that can be transferred to objects. Relying on gradient forces created by structured light, one can trap and move microscopic particles. Aside from the conservative action of gradient forces, light always pushes an object along its direction of propagation. Here, we demonstrate that gradientless light fields can exert pulling forces on arbitrary objects in a purely passive dielectric environment and without resorting to non-paraxial illumination, interference of multiple beams, gain or other exotic materials. The forces acting against the flow of light arise naturally due to the appropriate amplification of the photon linear momentum when light is scattered from one dielectric medium into another with higher refractive index. This situation opens up a number of intriguing prospects for optical forces and their effects on surface-bound objects. Here, we demonstrate that this new mechanism can be used to manipulate objects over macroscopic distances along dielectric interfaces.
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Laser heating of sulfuric acid droplets held in air by laser Raman tweezers

Oliver R. Hunt, Andrew D. Ward and Martin King
An optical trap is used to hold a droplet of concentrated sulfuric acid in the focus of Ar-ion laser (λ = 514.5 nm). The temperature and concentration of sulfuric acid in the droplet is calculated from the shifts and intensities of the Stokes-shifted Raman bands around 1000rel. cm−1. Aqueous sulfuric acid droplets can thus be used as a ‘thermometer’ for optically trapped droplets in air. It is demonstrated that the laser power can be kept low enough to prevent laser heating of the trapped droplet (necessary for hygrodynamic studies of atmospheric aerosols) and that high laser powers (in excess of 16mW) can result in heating of 5–10◦C.
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Sensing nanoparticles using a double nanohole optical trap

Abhay Kotnala, Damon DePaoli and Reuven Gordon
We use a double nanohole (DNH) optical trap to quantify the size and concentration of nanoparticles in solution. The time to trap shows a linear dependence with nanosphere size and a −2/3 power dependence with nanosphere concentration, which is in agreement with simple microfluidic considerations. The DNH approach has size-specificity on the order of a few nanometers, which was used to selectively quantify particles of a single size within a heterogeneous solution. By looking at individual trapping events, it is in principle possible to extend this approach to the ultimate limit of a single particle concentration, while also being able to operate at high concentrations in the same configuration. In addition, the DNH trap allows us to hold onto individual particles and thereby study constituents of a heterogeneous mixture. By repeating the trapping measurements on spherical particles of different refractive index, we found that the transmission step that indicates trapping scales empirically with the Clausius–Mossotti factor. This approach may be applied to several sensing applications, such as in the study of virus populations, where concentrations vary over many orders of magnitude.

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Transport and Trapping in Two-Dimensional Nanoscale Plasmonic Optical Lattice

Kuan-Yu Chen, An-Ting Lee, Chia-Chun Hung, Jer-Shing Huang, and Ya-Tang Yang
We report the transport and trapping behavior of 100 and 500 nm diameter nanospheres in a plasmon-enhanced two-dimensional optical lattice. An optical potential is created by a two-dimensional square lattice of gold nanostructures, illuminated by a Gaussian beam to excite plasmon resonance. The nanoparticles can be guided, trapped, and arranged using this optical potential. Stacking of 500 nm nanospheres into a predominantly hexagonal closed pack crystalline structure under such a potential is also reported.
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Two-Color Laser Printing of Individual Gold Nanorods

Jaekwon Do, Michael Fedoruk, Frank Jäckel, and Jochen Feldmann
We report on the deposition of individual gold nanorods from an optical trap using two different laser wavelengths. Laser light, not being resonant to the plasmon resonances of the nanorods, is used for stable trapping and in situ alignment of individual nanorods. Laser light, being resonant to the transversal mode of the nanorods, is used for depositing nanorods at desired locations. The power and polarization dependence of the process is investigated and discussed in terms of force balances between gradient and scattering forces, plasmonic heating, and rotational diffusion of the nanorods. This two-color approach enables faster printing than its one-color equivalent and provides control over the angular orientation (±16°) and location of the deposited nanorods at the single-nanorod level.

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Controllable Trapping of Nanowires Using a Symmetric Slot Waveguide

Nafiseh Zavareian and Reza Massudi
A controllable trapping method based on using a symmetric slot waveguide is proposed. The structure is composed of a subwavelength slot formed between two adjacent thin metallic films embedded in an infinite homogeneous dielectric medium. Generated near-field components interact with a nanowire and exert net force on it. Green’s function surface integral equation method is exploited for numerical calculation of the electric and the magnetic fields and, consequently, the radiation force acting on the nanowire. Casimir force is also obtained by calculating Maxwell stress tensor and using fluctuation–dissipation theorem. Results illustrate that depending on the width and the thickness of the slot, the radiation force and, consequently, the position of the stable equilibrium point change. By controlling the phase difference of the incident SPP waves it is possible to trap or release the nanowire at a specified position. In addition, results reveal that Casimir force moves nanowires toward the center of the slot and is maximum at the entrance of the slot with magnitude depending on the width and thickness of the slot.
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Localized cell stiffness measurement using axial movement of an optically trapped microparticle

Mary-Clare Dy; Shigehiko Kanaya; Tadao Sugiura

A simple optical tweezers design is proposed to manipulate particles in the axial direction and estimate particle position with nanometer sensitivity. Balb3T3 cell is probed using two different-sized particles, and the localized cell stiffness is evaluated using Hertz model. A series of experiments are performed to obtain the necessary parameters for the cell stiffness computation: particle displacement, trapping stiffness, force exertion, and cell deformation. The computed cell stiffness measurements are 17 and 40 Pa using 4 μm- and 2 μm-sized particles, respectively. Results suggest that the proposed optical tweezers scheme can measure the stiffness of a particular cell locale using Hertz model, offering insights about how cells respond to outside mechanical stimulus.

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Single crystal formation of amino acid with high temporal controllability by combining femtosecond and continuous wave laser trapping

Atsushi Miura, Yan-Hua Huang, Hiroshi Masuhara
We investigated laser trapping crystallization of glycine by using femtosecond (fs) laser as a trapping light source. Impulsively exerted fs laser pulses crystallized glycine more effectively than that induced by continuous wave (CW) laser trapping. Highly efficient crystallization and crystal growth behavior indicates fs laser irradiation increased the concentration not only at the focal spot, but also around the laser focus. Furthermore, we found that irradiation of fs pulses to CW laser-induced locally high supersaturation region enables immediate crystallization. Spatiotemporally controlled triggering of a single crystal formation with sub-second time resolution has achieved by integrating fs and CW laser trapping techniques.

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Single-cell optoporation and transfection using femtosecond laser and optical tweezers

Muhammad Waleed, Sun-Uk Hwang, Jung-Dae Kim, Irfan Shabbir, Sang-Mo Shin, and Yong-Gu Lee
In this paper, we demonstrate a new single-cell optoporation and transfection technique using a femtosecond Gaussian laser beam and optical tweezers. Tightly focused near-infrared (NIR) femtosecond laser pulse was employed to transiently perforate the cellular membrane at a single point in MCF-7 cancer cells. A distinct technique was developed by trapping the microparticle using optical tweezers to focus the femtosecond laser precisely on the cell membrane to puncture it. Subsequently, an external gene was introduced in the cell by trapping and inserting the same plasmid-coated microparticle into the optoporated cell using optical tweezers. Various experimental parameters such as femtosecond laser exposure power, exposure time, puncture hole size, exact focusing of the femtosecond laser on the cell membrane, and cell healing time were closely analyzed to create the optimal conditions for cell viability. Following the insertion of plasmid-coated microparticles in the cell, the targeted cells exhibited green fluorescent protein (GFP) under the fluorescent microscope, hence confirming successful transfection into the cell. This new optoporation and transfection technique maximizes the level of selectivity and control over the targeted cell, and this may be a breakthrough method through which to induce controllable genetic changes in the cell.
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Laser-induced breakdown of an optically trapped gold nanoparticle for single cell transfection

Yoshihiko Arita, Martin Ploschner, Maciej Antkowiak, Frank Gunn-Moore, and Kishan Dholakia

The cell selective introduction of therapeutic agents remains a challenging problem. Here we demonstrate spatially controlled cavitation instigated by laser-induced breakdown of an optically trapped single gold nanoparticle of diameter 100 nm. The energy breakdown threshold of the gold nanoparticle with a single nanosecond laser pulse at 532 nm is three orders of magnitude lower than water, which leads to nanocavitation allowing single cell transfection. We quantify the shear stress to cells from the expanding bubble and optimize the pressure to be in the range of 1–10 kPa for transfection. The method shows transfection of plasmid DNA into individual mammalian cells with an efficiency of 75%.
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Evaluation of single cell biomechanics as potential marker for oral squamous cell carcinomas: a pilot study

Janine Runge, Torsten E. Reichert, Anatol Fritsch, Josef Käs, Julia Bertolini, Torsten W. Remmerbach
Early detection of oral cancer is a major health issue. The objective of this pilot study was to analyze the deformability of healthy and cancer cells using a microfluidic optical stretcher (OS).
Different cancer cell lines, primary oral cancer cells and their healthy counterparts were cultivated and characterized respectively. A measurable deformation of the cells along the optical axis was detected, caused by surface stress, which is optically induced by the laser power.
All cells revealed a viscoelastic extension behavior and showed a characteristic deformation response under laser light exposure. The CAL-27/-33 cells exhibited the highest relative deformation. All other cells achieved similar values, but on a lower level. The cytoskeleton reacts sensitively of changing environmental conditions, which may be influenced by growth behavior of the cancer specimens. Nevertheless, the statistical analysis showed significant differences between healthy and cancer cells.
Generally malignant and benign cells showed significantly different mechanical behavior. Cancer related changes influence the composition of the cytoskeleton and thus affect the deformability, but this effect may be superimposed by cell cultivation conditions, or cell doubling time. These influences had to be substituted by brush biopsies to minimize confounders in pursuing investigations.
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Behavioral diversity in microbes and low-dimensional phenotypic spaces

David Jordan, Seppe Kuehn, Eleni Katifori, and Stanislas Leibler
Systematic studies of phenotypic diversity—required for understanding evolution—lag behind investigations of genetic diversity. Here we develop a quantitative approach to studying behavioral diversity, which we apply to swimming of the ciliate Tetrahymena. We measure the full-lifetime behavior of hundreds of individual organisms at high temporal resolution, over several generations and in diverse nutrient conditions. To characterize population diversity and temporal variability we introduce a unique statistical framework grounded in the notion of a phenotypic space of behaviors. We show that this space is effectively low dimensional with dimensions that correlate with a two-state “roaming and dwelling” model of swimming behavior. Temporal variability over the lifetime of an individual is correlated with the fraction of time spent roaming whereas diversity between individuals is correlated with the speed of roaming. Quantifying the dynamics of behavioral variation shows that behavior over the lifetime of an individual is strongly nonstationary. Analysis of behavioral dynamics between generations reveals complex patterns of behavioral heritability that point to the importance of considering correlations beyond mothers and daughters. Our description of a low-dimensional behavioral space should enable the systematic study of the evolutionary and ecological bases of phenotypic constraints. Future experimental and theoretical studies of behavioral diversity will have to account for the possibility of nonstationary and environmentally dependent behavioral dynamics that we observe.
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Laser-induced rotation and cooling of a trapped microgyroscope in vacuum

Yoshihiko Arita, Michael Mazilu & Kishan Dholakia
Quantum state preparation of mesoscopic objects is a powerful playground for the elucidation of many physical principles. The field of cavity optomechanics aims to create these states through laser cooling and by minimizing state decoherence. Here we demonstrate simultaneous optical trapping and rotation of a birefringent microparticle in vacuum using a circularly polarized trapping laser beam—a microgyroscope. We show stable rotation rates up to 5 MHz. Coupling between the rotational and translational degrees of freedom of the trapped microgyroscope leads to the observation of positional stabilization in effect cooling the particle to 40 K. We attribute this cooling to the interaction between the gyroscopic directional stabilization and the optical trapping field.

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