Optical traps provide tight confinement and very long storage times for atomic gases. Using a single focused beam from a CO₂ laser, we confine a mixture of spin-up and spin-down fermionic ⁶Li atoms, achieving storage times of ten minutes, and evaporative cooling to quantum degeneracy in seconds. A bias magnetic field tunes the gas to a collisional (Feshbach) resonance, producing extremely strong spin-pairing. This system now tests current many-body predictions for high-temperature superconductors, universal interactions in neutron stars, and hydrodynamic flow of quark-gluon plasmas, a state of matter that existed microseconds after the Big Bang.

We demonstrate the coherent transfer of the orbital angular momentum of a photon to an atom in quantized units of h, using a 2-photon stimulated Raman process with Laguerre-Gaussian beams to generate an atomic vortex state in a Bose-Einstein condensate of sodium atoms. We show that the process is coherent by creating superpositions of different vortex states, where the relative phase between the states is determined by the relative phases of the optical fields. Furthermore, we create vortices of charge 2 by transferring to each atom the orbital angular momentum of two photons. We subsequently use our technique to induce rotation of the condensate confined in an asymmetric, ring-shaped hybrid optical and magnetic trap. We observe the cloud rotating for up to 13 seconds, due to the superfluid character of the condensate.

Optical interference of two waves with different wave vectors generates a harmonic spatial profile of the optical intensity. This well known property combined with mechanical effect of light provides an excellent possibility to study the behavior of stochastic system in periodic potentials. Sub-micron size objects dispersed in liquid suffer from noticeable Brownian motion. This free motion can be significantly influenced (directed, suppress, modified) if the objects are illuminated by an optical interference pattern of sufficient intensity. Therefore, the Brownian dynamics in static periodic potentials of variable depth of the potential profile or extra tilt of the potential landscape can be studied. Moreover, applying the principle of optical conveyor combined with a precise method of object tracking with respect to the interference pattern paves the way for similar studies but in motional potential landscapes. We present a group of new experiments studying particle behavior in such fields.

Fluorescence correlation spectroscopy (FCS) was applied to investigate molecular translational diffusion in the solution of water, ethylene glycol, and heavy water under gradient light field of a near infrared (NIR) laser beam. The diffusion times of Rhodamine-6G in ethylene glycol and Rhodamine-123 in water became faster with an increase in the NIR laser power owing to absorption of the NIR light by the solvents. We also applied the radiation pressure of the NIR laser light to cadmium telluride (CdTe) nanoparticles dispersed in heavy water, resulting in increase in the average number of the CdTe particles in the confocal volume with increasing the NIR laser power.

In biological investigations, many protocols using optical trapping call for the possibility to trap a large number of particles simultaneously. Interference fringes provide a solution for massively parallel micro-manipulation of mesoscopic objects. Concurrently, the strength of traps can be improved by raising the slope of fringe profiles, such as to create intensity gradients much higher than the ones formed by sinusoidal fringes (Young's fringes). We use a multiple-beam interference system, derived from the classical Fizeau configuration, with semitransparent interfaces to generate walls of light with a very high intensity gradient (Tolansky fringes). These fringes are formed into a trapping set-up to produce new types of trapping templates. The possibility to build multiple trap arrays of various geometries is examined; a high number of particles can be trapped in those potential wells. The period of the fringes can easily be changed in order to fit traps sizes to the dimensions of the confined objects. This is achieved by modifying several parameters of the interferometer, such as the angle and/or the distance between the beam-splitter and the mirror. It is well known that optical trapping presents a great potential when used in conjunction with microfluidics for lab-on-a-chip applications. We present an original solution for multiple trapping integrated in a microfluidic device. This solution does not require high numerical aperture objectives.

Optical binding forces are present always when two and more colloidal particles are confined by external optical forces in a limited volume. They are caused by rescattering of the original incident field and they manifest themselves by a specific stationary displacements between confined micro-objects. Under certain circumstances the objects can self-organize into spatial arrangements creating so called optically bound matter. Detailed understanding of these forces might enable one to control this behavior. In this paper we present a theoretical study of the object self-organization in the field of two counter-propagating spatially incoherent non-diffracting beams. The field is uniform along the axis of propagation and so the studied phenomenon does not depend on the axial position of micro-objects. Therefore in this system the binding forces can be simply separated from the external forces of trapping field. We also compare this situation with setup of two counter-propagating Gaussian beams which are routinely used for studies of optical binding effects.

We investigate the extension of optical micromanipulation to integrated optics. In particular, we consider whether propagating light signals can cause mechanical reconfiguration of a device. While such forces are intrinsically weak, we predict theoretically that significant displacements can be achieved using various enhancement mechanisms. These include the use of high-index materials, high-Q (cavity quality factor) enhancement, and slow light in photonic crystals. Silicon optical waveguides have a considerable refractive index contrast with the surrounding air, with a ratio of roughly 3.45/1 at optical communications wavelengths. We show that the strong confinement of light to silicon magnifies optical forces arising from overlap in the guided modes of neighboring waveguides. Silica microsphere resonators are known to have extremely high cavity quality factors, in excess of 10⁸. We show that the quality factor of the resonator magnifies the optical force due to modal overlap between two neighboring spheres. Thirdly, we investigate slow-light enhancement of optical forces using photonic-crystal devices. We show that slow-light velocities give rise to larger forces for the same amount of signal power, enhancing optomechanical coupling effects. In addition to being of fundamental interest, our work suggests that optical manipulation may ultimately provide a route to all-optical conformational control and switching.

In the following paper we explore the dynamics of single colloidal particles and particle aggregates in a counterpropagating cavity-enhanced evanescent wave optical trap. For this study we make use of Fabry-Perot like cavity modes generated in a prism-coupled resonant dielectric waveguide. The advantage of using this type of optical structure is that there is an enhancement in the electric field of the evanescent at the sample surface that may be used to achieve greater coupling to colloidal particles for the purposes of optical micromanipulation. We demonstrate an order of magnitude increase in the optical forces acting on micrometer sized colloidal particles using cavity enhanced evanescent waves, compared with evanescent wave produced by conventional prism-coupling techniques. The combination of the enhanced optical interaction and the wide area illumination provided by the prism coupler makes it an ideal geometry for studying the collective dynamics of many particles over a large area. We study the different type of ordering observed when particles of different sizes are accumulated at the centre of this novel optical trap. We find that for large particles sizes (greater than 2μm), colloid dynamics are primarily driven by thermodynamics, whilst for smaller particles, in the range of 200-600nm, particles ordering is dictated by optical-matter interactions. We suggest a qualitative model for the observed optically induced ordering occurs and discuss how these results tie in with existing demonstrations of twodimensional optical binding.

In this paper, we report the excitation of whispering gallery modes (WGM) using a focused evanescent field. A focused evanescent field generated by total internal reflection at the cover glass-air interface was used to induce two-photon absorption in fluorescent polymer microcavities. By using a focused evanescent field it was possible to effectively couple light into the whispering gallery modes by the efficient overlap between the excitation volume and the cavity modes. The recorded whispering gallery mode spectra using an evanescent wave showed up to 38% enhancement in the spectral characteristics.

Two formulations of the Lorentz law of force in classical electrodynamics yield identical results for the total force (and total torque) of radiation on a solid object. The object may be surrounded by the free space or immersed in a transparent dielectric medium such as a liquid. We discuss the relation between these two formulations and extend the proof of their equivalence to the case of solid objects immersed in a transparent medium.

This paper examines two controversies arising within classical electromagnetism which are relevant to the optical trapping and micromanipulation community. First is the Abraham-Minkowski controversy, a debate relating to the form of the electromagnetic energy-momentum tensor in dielectric materials, with implications for the momentum of a photon in dielectric media. A wide range of alternatives exist, and experiments are frequently proposed to attempt to discriminate between them. We explain the resolution of this controversy and show that regardless of the electromagnetic energy-momentum tensor chosen, when material disturbances are also taken into account the predicted behaviour will always be the same. The second controversy, known as the plane wave angular momentum paradox, relates to the distribution of angular momentum within an electromagnetic wave. The two competing formulations are reviewed, and an experiment is discussed which is capable of distinguishing between the two.

With optical tweezer methods now firmly established and the nature of optical forces on individual particles well understood, one of the separate but related issues that has only recently come to the fore concerns the effects of intense optical radiation on inter-particle forces. It has already been established that such forces, which are not dependent on optical field gradients, can effect a weak binding between particles leading in some cases to optical clustering and in others to pattern formation. In this presentation it is shown by quantum electrodynamical analysis that a variety of other optomechanical effects can be produced in materials or systems subjected to the throughput of intense, non-resonant laser radiation. In particular, an optical electrostriction phenomenon is identified and shown to be widely operative in laser optical materials. Although a classical electrodynamical interpretation (in terms of interactions between induced dipoles) comfortably predicts the sign of the resulting mechanical force, it is shown that such a picture has significant limitations in addressing this fundamentally photonic phenomenon. The key parameters that determine the size and character of optical electrostriction are delineated and its significance is quantitatively assessed. The experimental challenges involved in characterizing such phenomena are also given a detailed appraisal.

Partial wave decomposition of incident beams is the first task to be performed to impose boundary conditions at the particle interface in the calculation of the scattering of spherical particles. The coordinate's origin must be in the center of the particle and not at high symmetry positions of the beam. This can be a quite complicated problem, especially when a full vectorial diffraction description of the electromagnetic fields and highly focused laser beams are required where the paraxial limit fails. Traditional approximation techniques have been used to proceed forward and to obtain numerical results. The main fault relies on a radial dependence of the beam shape coefficients, which limits the validity of such approximations. Here we prove that the radial dependence will emerge from the solid angle integration in this way obtaining an exact, closed expression, without any approximation, for the beam shape coefficients, for an arbitrary beam shape, origin and polarization, the special case of a Gaussian beam is presented.

Manipulation of micrometer sized particles with optical tweezers can be precisely modeled with electro dynamic theory using Mie's solution for spherical particles or the T-matrix method for more complex objects. We model optical tweezers for a wide range of parameters including size, relative refractive index and objective numerical aperture. We present the resulting landscapes of the trap stiffness and maximum applicable trapping force in the parameter space. These landscapes give a detailed insight into the requirements and possibilities of optical trapping and provide detailed information on trapping of nanometer sized particles or trapping of high index particles like diamond.

The ability to observe quantitatively mechanical events in real time of biological phenomena is an important contribution of the Optical Tweezers technique for life sciences. The measurements of any mechanical property involves force measurements, usually performed using a microsphere as the force transducer. This makes the understanding of the photonic force theory critical. Only very sensitive and precise experimental 3D photonic force measurements for any particle size will be able to discriminate between different theoretical models. In particular it is important to obtain the whole photonic force curve as a function of the beam position instead of isolate particular points. We used a dual trap in an upright standard optical microscope, one to keep the particle at the equilibrium position and the other to disturb it. With this system we have been able to obtain these force curves as a function of x, y and z position, incident beam polarization and wavelength. We investigated the optical forces for wavelengths in and out of Mie resonances of dielectric microspherical cavities for both TM and TE modes and compared the experimental results with the calculations performed with different models for the optical force.

The fluid lipid bilayer viscoelastic membrane of red blood cells (RBC) contains antigen glycolproteins and proteins which can interact with antibodies to cause cell agglutination. This is the basis of most of the immunohematologic tests in blood banks and the identification of the antibodies against the erythrocyte antigens is of fundamental importance for transfusional routines. The negative charges of the RBCs creates a repulsive electric (zeta) potential between the cells and prevents their aggregation in the blood stream. The first counterions cloud strongly binded moving together with the RBC is called the compact layer. This report proposes the use of a double optical tweezers for a new procedure for measuring: (1) the apparent membrane viscosity, (2) the cell adhesion, (3) the zeta potential and (4) the compact layer's size of the charges formed around the cell in the electrolytic solution. To measure the membrane viscosity we trapped silica beads strongly attached to agglutinated RBCs and measured the force to slide one RBC over the other as a function of the relative velocity. The RBC adhesion was measured by slowly displacing two RBCs apart until the disagglutination happens. The compact layer's size was measured using the force on the silica bead attached to a single RBC in response to an applied voltage and the zeta potential was obtained by measuring the terminal velocity after releasing the RBC from the optical trap at the last applied voltage. We believe that the methodology here proposed can improve the methods of diagnosis in blood banks.

In recent years there has been a growing interest in the use of optical manipulation techniques, such as optical tweezers, in biological research as the full potential of such applications are being realized. Biological research is developing towards the study of single entities to reveal new behaviors that cannot be discovered with more traditional ensemble techniques. To be able to study single cells we have developed a new method where a combination of micro-fluidics and optical tweezers was used. Micro-fluidic channels were fabricated using soft lithography. The channels consisted of a Y-shaped junction were two channels merged into one. By flowing different media in the two channels in laminar flow we were able to create a sharp concentration gradient at the junction. Single cells were trapped by the tweezers and the micro-fluidic system allowed fast environmental changes to be made for the cell in a reversible manner. The time required to change the surroundings of the cell was limited to how sharp mixing region the system could create, thus how far the cells had to be moved using the optical tweezers. With this new technique cellular response in single cells upon fast environmental changes could be investigated in real time. The cellular response was detected by monitoring variations in the cell by following the localization of fluorescently tagged proteins within the cell.

Living cells show a variety of morphological traits upon which numerous identification techniques have already been developed. However most of them involve lengthy biochemical procedures and can compromise the viability of the cell. We demonstrate a method to differentiate cells only on the basis of its trapping dynamics while it is being drawn into an optical trap (Optical Trapping Dynamics). Since it relies only on the inherent properties of the optical trap, without requiring external markers or biochemically sensitive spectroscopic techniques, it can be readily combined with existing optical tweezers setups. We applied it to the study of the yeast cell-cycle stages, showing, in particular, how it can be amenable for the measurement of the budding index of a cell population.

Light-scattering diagrams (phase functions) from single living cells and beads suspended in an optical trap were recorded with 30 ms time resolution. The intensity of the scattered light was recorded over an angular range of 0.5-179.5° using an optical setup based on an elliptical mirror and rotating aperture. Experiments revealed that light-scattering diagrams from biological cells exhibit significant and complex time dependence.

The way a cell reacts to a stimulus has a strong local nature based on the internal structure of it. Therefore models which describe, with a certain degree of precision, cell behaviours in response to a deformation of it as a whole cannot be extrapolated to the local response process. Under these assumptions the regional approach in single cell assays is earning more and more interest as it provides a more detailed insight on cells dynamics processes in terms of their morphology, and hence a more accurate description of the implied molecular entities. In the last decade, the development of a wide variety of optical trapping techniques has provided us a versatile tool to explore this locality of cells responses enabling a true "regional approach" and deepening our knowledge in the field. We here propose an apparatus based on multiple holographic optical tweezers and micro-stereolithography which allows an interactive control of the spatial-temporal characteristics of a trap pattern and the simultaneous application of different stimuli. These agents are kept separated from one another and from the cells via several custom-designed reservoirs fabricated via a micro-stereolithographic technique. It is worth noting that the work is more intended to propose a methodology or tool for meaningful assays rather than a new technique. As a consequence most of the efforts are being drawn towards to the simplification of the work flow to allow the device to actually be exploited and provide valuable data, and no more be a simple lab experiment.

Lipopolysaccharide (LPS) is one of the cell wall components of Gram-positive bacteria recognized by and interacted with receptor proteins such as CD14 on macrophage cells. Such a process plays an important role in our innate immune system. In this paper, we report the application of optical tweezers (λ = 1064nm Gaussian beam focused by a water-immersed objective lens with N.A. = 1.0) to the study of the dynamics of the binding of a LPS-coated polystyrene particle (diameter = 1.5μm) onto the plasma membrane of a macrophage cell. We demonstrated that the binding rate increased significantly when the macrophage cell was pre-treated with the extract of Reishi polysaccharides (EORP) which has been shown to enhance the cell surface expression of CD14 (receptor of LPS) on macrophage cells.

The combination of Raman spectroscopy and Optical Tweezers has been used to trap living cells and collect information about their biochemical state. Cells can continue living in such traps for periods of hours, allowing acquisition of time resolved Raman spectra. However no spatial information can be acquired as the cells continue to rotate and move in the single beam trap. Here we describe the development of Holographic Optical Tweezers (HOT) for the controlled movement of floating cells in order to construct their Raman images. Instead of a single trap, rapidly programmable multiple trapping points can be produced around the periphery of the cells to impede the rotational motion of the cell. By trapping and scanning the cell using HOT relative to a fixed Raman exciting laser, a point by point image of the cell can be constructed. We use an interactive program that permits us to position the trapping points relative to the live image feed we see from the microscope, using point and click. To demonstrate the possibilities of this technique images are shown of floating Jurkat cells.

Sperm cells from a domestic dog were treated with oxacarbocyanine DiOC₂(3), a ratiometrically-encoded membrane potential fluorescent probe in order to monitor the mitochondria stored in an individual sperm's midpiece. This dye normally emits a red fluorescence near 610 nm as well as a green fluorescence near 515 nm. The ratio of red to green fluorescence provides a substantially accurate and precise measurement of sperm midpiece membrane potential. A two-level computer system has been developed to quantify the motility and energetics of sperm using video rate tracking, automated laser trapping (done by the upper-level system) and fluorescent imaging (done by the lower-level system). The communication between these two systems is achieved by a networked gigabit TCP/IP cat5e crossover connection. This allows for the curvilinear velocity (VCL) and ratio of the red to green fluorescent images of individual sperm to be written to the hard drive at video rates. This two-level automatic system has increased experimental throughput over our previous single-level system (Mei et al., 2005) by an order of magnitude.

Central to the success of microfluidic systems has been the development of innovative methods for the manipulation of fluids within microchannels. We demonstrate a method for generating flow within a microfluidic channel using an optically driven pump. The pump consists of two counter rotating birefringent vaterite particles trapped within a microfluidic channel and driven using optical tweezers. The transfer of spin angular momentum from a circularly polarised laser beam rotates the particles at up to 10 Hz. We show the that the pump is able to displace fluid in microchannels, with flow rates of up to 200 μm³ s^-1 (200 fL s^-1). The direction of fluid pumping can be reversed by altering the sense of the rotation of the vaterite beads. We also incorporate a novel optical sensing method, based upon an additional probe particle, trapped within separate optical tweezers, enabling us to map the magnitude and direction of fluid flow within the channel. The techniques described in the paper have potential to be extended to drive an integrated lab-on-chip device, where pumping, flow measurement and optical sensing could all be achieved by structuring a single laser beam.

Photopolymerisation by computer controlled scanning of a focused laser beam is a powerful method to build structures of arbitrary complexity with submicrometer resolution. The procedure has already proven effective to produce complex structures that can be manipulated in optical tweezers. These micromechanical systems consist of static and moving parts and are expected to be building blocks of highly capable microfluidic systems. To enhance the efficiency of structure building, we developed single shot photopolymerisation. Instead of complicated multidimensional scanning the whole structure is generated simultaneously with special diffractive patterns. We experimented with fixed diffractive optical elements, kinoforms, and Spatial Light Modulators (SLMs). By using kinoforms, cross shaped structures were produced in single shots as an illustration. These propellers were produced about an order of magnitude faster than by simple scanning, and can be rotated by optical tweezer. The complexity of the structure depends on the quality of the kinoform and the available laser power. With the concerted movement of the appropriately chosen basic pattern and the sample, the building of more complicated structures can also be greatly accelerated due to the parallel nature of the polymerisation. The possibilities of photopolymerisation using SLM were also explored: the added flexibility using the programmable device is demonstrated.

Optical tweezers is a promising manipulation tool for objects in the range of micrometers to nanometers. Although there are many reported works on manipulating objects made of different materials and objects of irregular shapes, it is more suitable for non-opaque materials and objects that are symmetrical. Furthermore, there are potential damages on the objects arising from immense heat that is produced by the laser beam. These problems can be alleviated by trapping objects (micro-handles) and using them collectively as a gripper to indirectly hold and manipulate a target object. Holding denotes equilibrium of forces exerted by the tools on a target object. However, there still is a problem with this approach. When the trapping volume is larger than the size of a tool, target objects get pulled towards the center of the trapping volume. This breaks the force equilibrium and gripping thus fails. In this paper, we report a new design of tools that can overcome this problem. The tool is a slender object with one end acting as a probe while the other end is spherical so that trapping is easy. The length of the tool is designed to be larger than the radius of the trapping volume. Thus the target object is never pulled towards the trapping center. A group of multiple identical tools will surround and push a target object at the probe tips resulting in a stable grasp.

Optical tweezers use the electric-field gradient-force associated with tightly focused laser beams to trap micron-sized objects at the beam focus. Over the last few years optical tweezers have been revolutionized by the addition of spatial light modulators to split the laser beam into many traps that can be individually controlled; a technique called holographic optical tweezers. However, the reliance of optical tweezers on the gradient-force largely restricts their application to transparent objects that are not unduly sensitive to the effects of the laser light. Consequently, the manipulation of metallic particles or sensitive biomaterials can be problematic. In this work we use a holographic tweezers to position multiple silica beads acting as an optical gripper to lift, rotate and move micron-sized objects that otherwise do not lend themselves to tweezers control. We illustrate the use of the optical gripper under real-time joystick control to manipulate micron-sized metallic particles with nano-scale precision.

We investigate that components held in multiple optical traps can be manipulated and assembled together using snap-fit assembly. There are several works on manipulating microscopic objects with optical tweezers and assembling them. However, these techniques cannot sustain the assembled structure after turning off the laser source. In contrast, our technique utilizes snap-fit assembly so that assembled components do not detach. With this approach, components and sub-assemblies can be readily controlled in real-time and assembled into a permanent assembly. Our method can be used for constructing micrometer scale devices.

Electro-osmosis is an efficient means to move fluid in microfluidic channels. The flow is driven by the interaction of the electrical double layer at the channel wall with an electric field along the channel. The flow can be controlled by modifying the electrical &muparameters, either the charge of the channel wall or the electric field. If the surface chagre or the surface rsistance of the channel wall is sensitive to light, the flow can be modulated by light. We have demonstrated this effect by using photoconductive surfaces. The resistance change due to the illumination changes the electric field above the photoconductive layer and consequently changing the rate of fluid flow. By using channels where upon a photoresistive CdS surface a linear PDMS channel was placed, flow rate changes of an order of magnitude were achieved. This gives serious possibilities for optical control of flow. We further developed the method by building channel structures of more complicated patterns, e.g. Y-junctions. By appropriate illumination of the arms the flow direction could be selected between the arms optically. This unit is the basis of more complex flow patterns, it demonstrates the feasibility of optical control of such devices.

The application of laser radiation as a method for the manipulation of microscopic particle suspensions for biological, thermodynamic and microfluidic interests has brought about a revolution in micro-scale research across many different scientific disciplines. It has been shown that a diffraction limited focused laser can be used to trap microscopic particles whose refractive index is greater than their surrounding solvent. Termed Optical Trapping, work in this arena has yielded, new methods, techniques and applications that have flourished, and applications of this technology to areas of research involving microscopic systems for analysis, detection, separation and concentration have blossomed. A related technique, Optical Chromatography, used for particle separation involves loosely focusing a laser into a fluid flowing opposite to the direction of laser propagation. When microscopic particles in the flow path encounter this beam they are trapped axially along the beam and are pushed upstream from the laser focal point to rest at a point where the optical and fluid forces on the particle balance. Because optical and fluid forces are sensitive to differences in the physical and chemical properties of a particle, fine separations are possible. Recently, this method has been used to separate spores of different Bacillus species based on their optical and fluidic properties. We will describe how an optical chromatography beam directed into a tailored flow environment housed in a glass flowcell, has been adapted to operate as an optically tunable filter for the concentration or bioenrichment of colloidal and biological samples. Application of these methods and further design of fluidic and optical environments will allow for more specific identification, concentration and separation of many more microscopic particle and biological suspensions.

We demonstrate how the simple one-dimensional optical lattices can be used to sort micron-sized particles. Experimentally we focused on the simplest case where the periodical structure is created by an interference of two co-propagating beams and third non-interfering beam is used to compensate the radiation pressure caused by the previous two beams. We proved, that this geometry can be used for fast and efficient static sorting of microscopic objects without fluid flow.

We have developed an order of magnitude model for the the complete motion - translation and rotation - of spheroidal microparticles under the influence of intense optical landscapes and laminar flow using force and torque balances within the system. Our fields of interest are periodic interference profiles, sometimes termed optical landscapes, that can be formed by simple holography. When given an arbitrary landscape, our model predicts that, in general, spheroidal particles become trapped at a lower potential threshold than do spherical particles of an equivalent volume. In addition, we show that optical landscapes exhibit exponential trapping selectivity based on particle size and shape, effectively adding a further dimension of control over which to trap, influence, and sort particles within the same flow.

We demonstrate the sequential spatial separation of a solution consisting of a mixture of two microspheres with different diameters using a dynamic optical interferometery scheme. Two coherent lasers beams are focused together through an objective lens to form an in-plane standing wave. By linearly increasing the phase of one of incoming beams relative to the other, the optical lattice is translated. The optical forces on particles with different sizes depends on the spacing of the standing wave relative to the particle diameter; therefore, by adjusting the spacing of the standing wave so as to minimize the interaction of particles of one size with the optical lattice, all other particles can be swept out by the translating potential wells that are associated with the intensity maxima of the standing wave, while the selected particles remain trapped in the overall center of the Gaussian beam envelope of the optical lattice. Here, we demonstrate the selectivity of this optical conveyor belt by dragging smaller particles out to one side of an ensemble while simultaneously keeping the larger ones trapped. The Brownian dynamics of particles translated in an optical lattice and measurements of the associated optical force are also presented.

We present a detailed theoretical discussion and experimental analysis of an interferometric optical trapping device that allows efficient sorting of particles, including biological samples, either by size or refractive index. This technique involves no microfluidic flow, but it is based on the specific response of different microparticles to an interference pattern of fringes vibrating with a periodic but non-symmetric time modulation function. The performance of the system is analyzed in terms of the different control parameters, such as the period of the fringes, the vibration amplitude and frequency, and the power level in the sample. We discuss the possibility of using this system to characterize unknown samples.

Optoelectronic tweezers (OET) provides a non-invasive, low-power, optical manipulation tool for trapping, transporting, and separating microparticles, cells, and other bioparticles. The OET device uses a photosensitive layer to form "virtual electrodes" upon exposure to light, creating non-uniformities in an applied electric field. The electric field gives rise to a force known as dielectrophoresis: microparticles move as a result of the non-uniformities in the electric field imparting unequal forces on the induced dipoles of the particles. These virtual electrodes can be actuated with low optical intensities, enabling the use of incoherent light sources and direct imaging techniques to create optical manipulation patterns in real-time. In this paper, we demonstrate OET operation on live cells, including the trapping and manipulation of red and white blood cells, and the automated collection of HeLa cells. Automated size-based sorting is performed on a mixture of 15- and 20-μm-diameter polystyrene beads, and dielectric property-based separation is used to differentiate between live and dead white blood cells.

We present a study into the small particle size and resolution limits of Light Induced Dielectrophoresis (LIDEP). Here the illumination of a photoconductive layer creates virtual electrodes whose associated electric field gradients cause the dielectrophoretic response of the particles. In this way a potential energy landscape can be created that is optically controlled giving reconfigurable control over a large area [1]. In this paper we discuss the interlinked limits of size of particle it is possible to manipulate and the resolution these particles can be manipulated with. We compare traditional dielectrophoresis (DEP) experiments with LIDEP experiments, and discuss the mechanisms behind the physical limits comparing the effects of carrier diffusion verses the spreading of the electric fields in the medium.

Particles that can be trapped in optical tweezers range in size from tens of nanometres to tens of micrometres. Notably, this size range includes large single molecules. We show experimentally, in agreement with theoretical expectations, that optical tweezers can be used to manipulate single molecules of polyethylene oxide suspended in water. The trapped molecules accumulate without aggregating, so the optical trap offers a method of controlling the concentration of macromolecules in solution. Potential applications are the micromanipulation of nanoparticles, nanoassembly, microchemistry, and the study of biological macromolecules.

We describe two methods to optically measure the torque applied by the orbital angular momentum of the trapping beam in an optical tweezers setup. The first decomposes the beam into orbital angular momentum carrying modes and measures the power in each mode to determine the change in angular momentum of the beam. The second method is based on a measuring the torque transfer due to spin angular momentum and the linear relationship between rotation rate and applied torque to determine the orbital angular momentum transfer. The second method is applied to measuring the transfer efficency for different particle-mode combinations. We present the results of these experiments and discuss some of the difficulties encountered.

A modulated laser beam by a phase pattern exp(ilθ) can be focused by an objective into a ring-like optical vortex, where l is a constant and θ is the azimuth angle. The vortex is capable of trapping the particles nearby and circulating them along the ring. This phenomenon is often explained involving Fourier optics and the transfer of orbital angular momentum (OAM). Although Fourier optics transforms the electric field distribution of the modulated laser beam behind the phase pattern to that of the vortex, it does not include both the path and OAM of the photons of the electromagnetic wave. Therefore, it is difficult to further trace the transfer of OAM from the photons to the particles in the vortex. In this paper, we propose a simple and intuitive view to the origin of optical vortex. By analyzing the relationship of the intensity distributions between the phase of the phase pattern and the intensity of the vortex by utilizing Fourier transform, we propose that the phenomenon of vortex also involve the transfer of linear momentum on the vortex plane transversely.

The central maximum of a Bessel beam offers a "non-diffracting" focal line of light that is useful in the fields of optical trapping and micromanipulation. This paper discusses the design and performance of diffractive optics for converting a Gaussian beam into a Bessel beam. The theoretical foundation of Bessel beams will be reviewed along with their optical properties. Bessel beams provide several unique characteristics such as a large depth of field and self-reconstruction. It is well known that the depth of field of a Bessel beam is larger than that of a Gaussian beam of equivalent size. However, this comes at the expense of very little power contained within the central maximum of the Bessel beam. Optical modeling and beam propagation methods are used to analyze what effect the number of rings has on the depth of field. This is an important consideration if Bessel beams are ever to be used in the fields of optical interconnects and imaging or in the area of laser processing. Where appropriate, comparisons are made between Bessel and Gaussian beams.

Aerosol droplets are guided over mm distances using single beam optical traps. The micron-sized particles are confined in two dimensions and guided along the direction of beam propagation. Both Gaussian and Bessel beam geometries are compared for water, ethanol and dodecane droplets. The observed trapping of multiple droplets in 1-D arrays will also be discussed.

The Tunable Acoustic Gradient Index of Refraction (TAG) lens is shown to be an alternative method of generating Bessel beam for optical manipulation. The TAG lens exhibits a combination of tunability, optical throughput, and fast switching speed, overcoming many limitations linked with conventional Bessel beam techniques. A refractive fluid is contained within a circular piezoelectric that is driven with an AC signal to establish a periodic index of refraction in the liquid. Simple changes in amplitude or frequency of the driving signal allows for the rapid modification to the transmitted pattern. The switching speed is characterized by the time to reach a steady state pattern, which is shown to depend primarily on the viscosity of the filling liquid. Times between 300 and 2000 μs are obtained corresponding to fluids with viscosity of 640 cs and 0.65 cs respectively.

We demonstrate the use of supercontinuum radiation to provide enhanced guiding distances of microscopic particles compared to the standard continuous wave or femtosecond lasers. Our technique relies on the chromatic aberration of the lens used to form an elongated focal region within which guiding takes place. The resulting beam profile has been modelled and shows that for a Gaussian input beam, the intensity profile after the lens can be considered as a sum of Gaussians, one for each wavelength but with varying focal position due to dispersion. Our experimental investigations compare radiation from continuous wave (bandwidth <1nm) and femtosecond pulsed (bandwidth > 100nm) lasers as well as supercontinuum radiation (bandwidth > 450nm) and show good agreement with theory.

White light supercontinuum, which is generated by coupling short laser pulses into a nonlinear photonic crystal fiber, not only covers an extremely broad wavelength range (e.g., from visible to near infrared) but also has high spatial coherence. As a result, tightly focused supercontinuum can be used to trap a single particle and simultaneously to perform broad-band ultra-sensitive optical spectroscopy at a single particle level. In this paper we investigate the optical scattering spectroscopy of a single particle in white light supercontinuum optical tweezers. Lorenz-Mie theory and Fourier angular spectrum analysis are used to model the scattering of tightly focused supercontinuum by a uniform spherical scatterer. In addition, Born approximation method is applied to analyze scattering by non-spherical weak scatterers. Unlike conventional ensemble averaged spectroscopy, single particle spectroscopy has the unique capability to probe the properties of individual particles, which can lead to many important applications such as ultrasensitive sensing and nanoparticle characterization.

We demonstrate the use of holographic optical tweezers for the optical trapping and manipulation of arrays of airborne water droplets (aerosols). Making use of a phase-only spatial light modulator we present evidence of stable, interactive manipulation of both single and multiple aerosol droplets, of the order of 10 microns in diameter, and also their controlled coagulation. We discuss the advantages, disadvantages, and limitations of using a spatial light modulator for droplet manipulation including the implications of the update speed of the device (a Holoeye LC-R2500 SLM), diffraction efficiency, and droplet growth and evaporation due to laser intensity variations. We will examine the generic difficulties of trapping in air, working in the absence of inertial damping. Finally we will discuss the applications of the above work in fields such as atmospheric chemistry and microfluidic microchemical reactors whilst presenting preliminary results on fusion of two or more droplets of differing phases.

We demonstrate a technique for the multi-point measurement of fluid flow in microscopic geometries. The technique consists of an array of microprobes can be simultaneously trapped and used to map out the fluid flow in a microfluidic device. The optical traps are alternately turned on and off such that the probe particles are displaced by the flow of the surrounding fluid and then re-trapped. The particles' displacements are monitored by digital video microscopy and directly converted into velocity field values. The techniques described have potential to be extended to drive an integrated lab-on-chip device, where pumping, flow measurement and optical sensing could all be achieved by structuring a single laser beam.

The shapes of unilamellar lipid vesicles are driven out of equilibrium by direct forcing with holographic optical tweezers. Vesicles have been studied extensively due to their relevance as a model for the membrane of cells as well as their potential practical uses e.g. for drug delivery or chemical confinement. We use multipoint laser tweezers formed by a spatial light modulator (holographic optical tweezers) to apply forces to such vesicles in several points simultaneously. To apply forces we utilize an index of refraction difference between the fluid inside the vesicle and the external fluid. Since this higher index of refraction material is fluid, the vesicle shape can changes in response to the optical forces. This shape change reveals the mechanical properties of vesicles subject to multiple stresses. We find that the surface forces on the membrane are localized near the points of forcing. Restoring forces from lipid tethers are used to estimate the total applied optical forces, which are below the pN level. The relaxation of deformations can be decomposed into its Fourier modes. The relaxation of all observable modes can be described well by a third order Landau equation. Ellipsoidal deformations relax more slowly than higher order deformation modes.

We present and discuss a new experimental setup to perform small angle X-ray scattering and diffraction (SAXSD) of localized liposome colloidal microparticles. A home-built inverted infrared laser tweezers microscope is used to trap, manipulate and aggregate micron-scale liposome particles at single locations inside a 100 microns glass capillary. The micro-focused X-ray and the laser beams are aligned to intersect each other perpendiculary, allowing to associate the X-ray diffraction signal to the micron-sized region of interest inside the capillary. Throughout the laser tweezer setup, using diffractive optical elements implemented on a spatial light modulator, we are able to manipulate small aggregates of colloidal particles (liposomes) and fix them in the optical path of the X-ray beam. We present and discuss first scattering and diffraction experiments on phospholipid liposomes, at the ID13 microfocus beamline of the European Synchrotron Radiation Facility (ESRF). The results demonstrate that we can push the limit of measurable cluster size close to a single liposome.

Optical technology is rapidly finding novel applications in several exiting bioanalytical, biological, and biomedical applications. Optical beams are increasingly used for bio-fluidic sample manipulation in BioMEMS devices replacing convectional mechanical, electrostatic, and electrokinetic methods. This paper presents novel multiphysics computational approach for modeling optical interaction with fluidic, thermal, mechanical, and biological processes. We present a model of optical manipulation of particles and biological cells with laser beams. Computational results are compared to available experimental data from laboratory experiments and from practical engineered optical bio microdevices. The modeling approach is demonstrated on selected specific applications of optical manipulation of micro spheres, micro cylinders, and optical manipulation and sorting of biological cells in microfluidic cytometers.

Because of its non-invasive nature, optical tweezers have emerged as a popular tool for the studies of complex fluids and biological cells and tissues. The capabilities of optical tweezer-based experimental instruments continue to evolve for better and broader applications, through new apparatus designs and integrations with microscopic imaging techniques. In this paper, we present the design, calibration and applications of a powerful microrheometer that integrates a novel high temporal and spatial resolution dual-beam oscillating optical tweezer-based cytorheometer (DOOTC) with spinning disk confocal microscopy. The oscillating scheme detects the position of micron-size probe particles via a phase-sensitive lock-in amplifier to greatly enhance sensitivity. The dual-beam scheme ensures that the cytorheometer is insensitive to sample specimen background parameter variances, and thus enables the investigation of micromechanical properties of biological samples, which are intrinsically inhomogeneous. The cytorheometer system is demonstrated to be capable of measuring dynamic local mechanical moduli in the frequency range of 0.1-150 Hz at up to 2 data point per second and with nanometer spatial resolutions, while visualizing and monitoring structural properties in situ. We report the results of system applications in the studies of bovine skin gelatin gel, purified microtubule assemblies, and human alveolar epithelial cells. The time evolution of the storage moduli G' and the loss moduli G'' of the gel is recorded for undisturbed gel-forming process with high temporal resolution. The micromechanical modulus G* of polymerized microtubule network as a function of frequency are shown to be both inhomogeneous and anisotropic consistent with local structures revealed by confocal imaging. The mechanical properties of A549 human lung cells as a function of temperature will be reported showing significant decrease in cell stiffness at higher temperature.

The length and time scales accessible to optical tweezers make them an ideal tool for the examination of colloidal systems. Embedded high-refractive-index tracer particles in an index-matched hard sphere suspension provide 'handles' within the system to investigate the mechanical behaviour. Passive observations of the motion of a single probe particle give information about the linear response behaviour of the system, which can be linked to the macroscopic frequency-dependent viscous and elastic moduli of the suspension. Separate 'dragging' experiments allow observation of a sample's nonlinear response to an applied stress on a particle-by particle basis. Optical force measurements have given new data about the dynamics of phase transitions and particle interactions; an example in this study is the transition from liquid-like to solid-like behaviour, and the emergence of a yield stress and other effects attributable to nearest-neighbour caging effects. The forces needed to break such cages and the frequency of these cage breaking events are investigated in detail for systems close to the glass transition.

By performing experiments at an air-water interface, we operate Holographic Optical Tweezers in a qualitatively new environment. In this regime, trapping and moving of micro particles may allow access to parameters like local viscosity and surface tension. Polystyrene micro beads are naturally stabilized in the interface due to a minimum in surface energy. For this reason, they can also be manipulated by light patterns with small axial field gradients, without causing the particles to escape due to scattering forces. In this manner, the interface provides a true two-dimensional "working environment", where particles can be manipulated with high effciency. For example, we demonstrate different optical "micro tools", which utilize scattering and gradient forces to enable controlled transport of matter within the surface.

Optical tweezers enable non-destructive, contact-free manipulation of ultrasound contrast agent (UCA) microbubbles, which are used in medical imaging for enhancing the echogenicity of the blood pool and to quantify organ perfusion. The understanding of the fundamental dynamics of ultrasound-driven contrast agent microbubbles is a first step for exploiting their acoustical properties and to develop new diagnostic and therapeutic applications. In this respect, optical tweezers can be used to study UCA microbubbles under controlled and repeatable conditions, by positioning them away from interfaces and from neighboring bubbles. In addition, a high-speed imaging system is required to record the dynamics of UCA microbubbles in ultrasound, as their oscillations occur on the nanoseconds timescale. In this work, we demonstrate the use of an optical tweezers system combined with a high-speed camera capable of 128-frame recordings at up to 25 million frames per second (Mfps), for the study of individual UCA microbubble dynamics as a function of the distance from solid interfaces.

We suggest a novel 3-D probing technique for measuring micro parts which have high aspect ratio such as a groove or a deep-hole. This technique uses the optical force exerted on a dielectric microsphere at the tip of optical fiber so called the fiber optical trapping. A microsphere is trapped at the tip of an optical fiber which has micrometer size of the diameter and sub micrometer size of the tapered tip. The optical source is a Nd:YAG laser with the wavelength of 1064 nm. A fiber optical trapping system is quite similar to a conventional CMM. The roll of a microsphere is the same as the probe sphere of a conventional CMM and an optical fiber works as a stylus shaft. The micro optical fiber part is thinner than the diameter of the microsphere and longer than the depth of shapes such as deep holes and grooves, which enable to make an approach to a steep angle surface of a work piece with high aspect ratio.

In the area of manipulating microscopic biological specimen, optical trapping has proven its worth. Still, many potential microbiological applications can benefit when the experiment is assisted by a computer and capable of running either with only limited supervision or full automation. Here we have implemented the Generalized Phase Contrast (GPC) method of optical trapping in a microfluidic system, and show how an experiment can be easily made to run autonomously, while the computer continuously adapts the light pattern to trap yeast cells passing through the trapping volume. The optical trapping takes place in a microfluidic system where two channels meet, allowing for separate injection of specimen and work media. Yeast cells are trapped near the surface of the microchannel at flow rates that give particle speeds of more than 50 μm/s. We demonstrate the ability of GPC-based traps to hold the cells in specific positions and observe the displacement of the cells from respective trap centers. Finally we exploit the speed of the GPC system by dynamically detect yeast cells using a CCD camera and immediately create traps at their positions at flow rates that exceed what a human operator would be able to handle. The optical system was found to be easily expanded and the attention could be kept on maintaining optimal conditions for the yeast.

We create long polymer nanotubes by directly pulling on the membrane of polymersomes using either optical tweezers or a micropipette. The polymersomes are composed of amphiphilic diblock copolymers and the nanotubes formed have an aqueous core connected to the aqueous interior of the polymersome. We stabilize the pulled nanotubes by subsequent chemical cross-linking. The cross-linked nanotubes are extremely robust and can be moved to another medium for use elsewhere. We demonstrate the ability to form networks of polymer nanotubes and polymersomes by optical manipulation. The aqueous core of the polymer nanotubes together with their robust character makes them interesting candidates for nanofluidics and other applications in biotechnology.

Instrumentation and methodologies for single molecule force spectroscopy on bacterial adhesion organelles by the use of force measuring optical tweezers have been developed. A thorough study of the biomechanical properties of fimbrial adhesion organelles expressed by uropathogenic E. coli, so-called pili, is presented. Steady-state as well as dynamic force measurements on P pili, expressed by E. coli causing pyelonephritis, have revealed, among other things, various unfolding and refolding properties of the helical structure of P pili, the PapA rod. Based on these properties an energy landscape model has been constructed by which specific biophysical properties of the PapA rod have been extracted, e.g. the number of subunits, the length of a single pilus, bond lengths and activation energies for bond opening and closure. Moreover, long time repetitive measurements have shown that the rod can be unfolded and refolded repetitive times without losing its intrinsic properties. These properties are believed to be of importance for the bacteria's ability to maintain close contact with host cells during initial infections. The results presented are considered to be of importance for the field of biopolymers in general and the development of new pharmaceuticals towards urinary tract infections in particular. The results show furthermore that the methodology can be used to gain knowledge of the intrinsic biomechanical function of adhesion organelles. The instrumentation is currently used for characterization of type 1 pili, expressed by E. coli causing cystitis, i.e. infections in the bladder. The first force spectrometry investigations of these pili will be presented.

With the evolution of single molecule techniques as force-scope optical tweezers, it has become possible to perform very accurate measurements of the elastic properties of biopolymers as e.g. DNA. Nucleic acid elasticity is important in the interaction of these molecules with proteins and protein complexes in the living cell. Most experimental and theoretical effort has been aimed at uncovering and understanding of the behavior of polymers with contour lengths significantly longer than their persistence length. The well-established Worm-Like-Chain model has been modified such that a satisfactory description of such long biopolymers is available. However, in many single molecule experiments, such as the unfolding of RNA stem-loops¹ and RNA pseudoknots,² one is dealing with biopolymers whose contour lengths are comparable to persistence lengths. A full understanding of such curves requires an understanding of the physics of short biopolymers. For such cases, theories are just beginning to emerge and there is hardly any experimental data available. We target this problem by optical tweezers quantitative force-extension measurements on short biopolymers. The biopolymers used are primarily double stranded DNA whose total length ( 300 nm) is comparable to their persistence length ( 50 nm). As a control of our equipment and methods, we also stretch longer dsDNA (1100 nm), the force-extension curves of which resemble those in literature.³ For the short DNA the force-extension curves qualitatively resemble those predicted by WLC theories, but a reasonable fit can only be made if the persistence length is allowed to be a fitting parameter. If made a fitting parameter, the 'apparent persistence length' is found as 8.7±4 nm, a number which is significantly lower than the real physical value.

The bacteriophage φ29 portal motor is capable of packaging the φ29, 19.3 Kbp, genome to high density into its preformed capsid. The packaging process must overcome the forces due to confining the highly negative charge of the DNA to a small volume, as well as the forces due to bending the DNA on length scales smaller than one persistence length. Both of these energetic considerations can be modulated by the ionic nature of the buffer DNA packaging occurs in. To measure the effects of DNA charge shielding on the packaging process, we studied the dynamics of DNA packaging by optical tweezers in a variety of different ionic conditions. We looked at the effects monovalent, divalent, and trivalent cations have on the motor function and its dependence on external force and, we observed the rate of DNA packaging at nominal force as a function of capsid filling. Specifically, we varied the concentrations of Na⁺, Mg⁺², and cobalt hexamine in the solution bathing the bacteriophage during packaging to see what effects, if any, these cations have. From these measurements, we present an inferred internal force as a function of percent filling of the bacteriophage capsid in a variety of ionic environments. Preliminary analysis suggests the ionic environment can modulate internal pressure, with the presence of higher valence cations better shielding the packaged DNA resulting in lower internal pressures.

A key step in the life cycle of many viruses, including bacteriophages, adenoviruses, and herpesviruses, is the packaging of replicated viral genomes into pre-assembled proheads by the action of ATP-dependent portal motor complexes. Here we present a method that allows the initiation of packaging by single complexes to be studied using optical tweezers. A procedure is developed for assembling phage Φ29 prohead-motor complexes, which are demonstrated to bind and begin translocation of a target DNA molecule within only a few seconds. We show that the Φ29 DNA terminal protein (gene product 3), which functions to prime DNA replication, also has a dramatic effect on packaging. The DNA tether length measured immediately after binding varied from ~30-100% of the full length, yet shortened monotonically, indicating that packaging does not strictly begin at the terminal end of the DNA. Removal of the terminal protein eliminated this variability, causing packaging to initiate at or very near the end of the DNA. These findings, taken together with electron microscopy data, suggest that rather than simply threading into the portal, the motor captures and dynamically tensions a DNA loop, and that the function of the terminal protein is to load DNA segments on both sides of the loop junction onto separate DNA translocating units.

Optical tweezers have broad applications in studies of structures and processes in molecular and cellular biophysics. Use of optical tweezers for quantitative molecular-scale measurement requires careful calibration in physical units. Here we show that DNA molecules may be used as metrology standards for force and length measurements. Analysis of DNA molecules of two specific lengths allows simultaneous determination of all essential measurement parameters. We validate this "biological calibration" method experimentally and with simulated data, and show that precisions in determining length scale factor (~0.2%), length offset (~0.03%), force scale factor (~2%), and compliance of the traps (~3%) are limited only by current measurement variation, much of which arises from polydispersity of the microspheres (~2%). We find this procedure to be simpler and more convenient than previous methods, and suggest that it provides an easily replicated standard that can insure uniformity of measurements made in different laboratories.

Looping and cleavage of single DNA molecules by the two-site restriction endonuclease Sau3AI were measured with optical tweezers. A DNA template containing many recognition sites was used, permitting loop sizes from ~10 to 10,000 basepairs. At high enzyme concentration cleavage events were detected within 5 seconds and nearly all molecules were cleaved within 5 minutes. Activity decreased ~10-fold as the DNA tension was increased from 0.03 to 0.7 pN. Substituting Ca²⁺ for Mg²⁺ blocked cleavage, permitting measurement of stable loops. At low tension, the initial rates of cleavage and looping were similar (~0.025 s^-1 at 0.1 pN), suggesting that looping is rate limiting. Short loops formed more rapidly than long loops. The optimum size decreased from ~250 to 45 bp and the average number of loops (in 1 minute) from 4.2 to 0.75 as tension was increased from 0.03 to 0.7 pN. No looping was detected at 5 pN. These findings are in qualitative agreement with recent theoretical predictions considering only DNA mechanics, but we observed weaker suppression with tension and smaller loop sizes. Our results suggest that the span and elasticity of the protein complex and protein-induced DNA bending and wrapping play an important role.

This paper presents a novel method for manipulating single chromosomal DNA, which is intended for the use in highresolution genomic studies. Such operations as translocation, winding and unwinding of single DNA fiber are achieved using optically-driven micro-fabricated structures, including micro-hooks and micro-bobbins for picking-up and winding DNA, with a typical dimension of several μm. The geometry of the laser-manipulated micro-structures is designed in such a way that a spontaneous orientation occurs with its major axis parallel to the laser beam and accepts a DNA fiber. While monitoring under a fluorescence microscope, yeast chromosomal DNA is first extended to the full length by electroosmotic flow. Then the micro-hooks are dispensed in the solution, and a DNA fiber is picked up with the microhook which is driven by a focused laser beam, to separate the targeted DNA from the others. The winding is achieved with a pair of micro-bobbins. The laser is split into two, the first beam being fixed, and the second movable circularly around the first. When the bobbins are made into contact with DNA and revolving motion started, the fiber is wound and suspended between them. The unwinding can be achieved just by reversing the revolving motion.

We present a new and precise method how to employ a periodic interference illumination for axial particle position determination. Particle movement with respect to the interference structure of illumination is followed by changes in the light field scattered by the particle. Analyses of these changes together with their calibration provide an excellent way how to determine not only the particle position with respect to the camera but especially with respect to the illuminating field structure. The algorithm was used in a standing wave optical trap for determination of the trap properties and particle behavior even in the standing wave in motion.

We demonstrate an example of 'confocal-tweezers' wherein confocal images and precise optical force measurements, using photodiodes, are obtained simultaneously in the x-y plane without moving the objective lens. The optical trap is produced using a 1.064μm cw laser and is combined with Leica's TCS SP5 broadband confocal microscope to trap and image living cells. The unique method by which the confocal images are created facilitates the acquisition of images in areas far from the trapping location. In addition, because the scanning process involves moving galvanic mirrors independently of the objective, the trap is held stable in position and is not subject to any error in position for the x-y scan. We have successfully trapped and confocally imaged 80nm gold colloids, 150nm gold colloids and 1μm polystyrene beads whilst making quantitative measurements of the force applied by the trap on each bead. To the best of our knowledge this is the first time that anyone has combined precise force measuring optical tweezers with confocal microscopy. We also discuss some of the technical challenges involved in advancing the experimental set up to make quantitative force measurements in combination with 3D stacking. Having proven the potential of this system in 2D, we hope to develop it further to investigate the nano-mechanics of cell division through the attachment of gold beads to fluorescently labelled organelles in S. pombe yeast cells.

One of the most promising ways to study the biochemistry of single floating cells is to combine the techniques of optical tweezers and Raman spectroscopy (OTRS). This can reveal the information that is lost when ensemble averages are made over cell populations, like in biochemical assays. However, the interpretation of the acquired data is often ambiguous. Indeed, the trapped living cell continues to move and rotate in the optical trap not only because of the Brownian motion, but also because of its inherent biological motility and the variation of its shape and size. This affects both Rayleigh and Raman light scattering. We propose the use of Rayleigh scattering to monitor the growth of a single optically trapped yeast cell, while OTRS measurements are being performed. For this purpose, we added a quadrant photodiode to our OTRS setup. The cell orientation in the optical trap is shown to vary as the cell growth proceeds, especially when it becomes asymmetrical (budding) or it changes its size or shape considerably (living and growing cell). Control experiments, performed using heat-treated cells and polystyrene beads, confirm that this behavior is a consequence of the cell growth. These measurements have to be taken into account in the interpretation of Raman spectra so as not to incorrectly attribute variations in the spectra to change in the biochemical constituents of the cell if they are in fact due to a change of the orientation of the cell in the trap.

The displacements of a dielectric microspheres trapped by an optical tweezers (OT) can be used as a force transducer for mechanical measurements in life sciences. This system can measure forces on the 50 femto Newtons to 200 pico Newtons range, of the same order of magnitude of a typical forces induced by flagellar motion. The process in which living microorganisms search for food and run away from poison chemicals is known is chemotaxy. Optical tweezers can be used to obtain a better understanding of chemotaxy by observing the force response of the microorganism when placed in a gradient of attractors and or repelling chemicals. This report shows such observations for the protozoa Leishmania amazomenzis, responsible for the leishmaniasis, a serious tropical disease. We used a quadrant detector to monitor the movement of the protozoa for different chemicals gradient. This way we have been able to observe both the force strength and its directionality. The characterization of the chemotaxis of these parasites can help to understand the infection mechanics and improve the diagnosis and the treatments employed for this disease.

Recent work has indicated the potential of light to guide the growth cones of neuronal cells using a Ti:Sapphire laser at 800 nm (Ehrlicher et al, PNAS, 2002). We have developed an optical set-up that has allowed, for the first time, the direct comparison of this process at near infrared wavelengths. A high number of growth cones were studied in order to provide a detailed statistical analysis. Actively extending growth cones of the neuroblastoma cell-line, NG108, can be guided at not only 780 nm, but also at 1064 nm. These wavelengths are an appropriate choice for guidance experiments, as wavelengths in the visible spectrum and UV are highly absorbing by cells and lead to death by phototoxicity and thermal stress. At 780 nm, 47% of actively extending growth cones were found to turn towards the focused incident light by at least 30° (n=32 growth cones). At 1064 nm, 61% of cells were successfully guided (n=31 growth cones). This suggests that the light detection mechanism within the cell is not due a single protein with a defined activity wavelength as occurs for example with the photoreceptor family of opsin proteins in the mammalian eye. We present two novel mechanisms of light induced neuronal guidance which are not related to temperature increases, or optical tweezing of the growth cone. We are also now identifying the signaling pathways that mediate this phenomenon.

In previous studies we have shown that the second harmonic 532 nm, from a picosecond frequency doubled Nd:YAG laser, can cleanly and selectively disrupt spindle fiber microtubules in live cells (Botvinick et al 2004, Biophys. J. 87:4303-4212). In the present study we have ablated different locations and amounts of the metaphase mitotic spindle, and followed the cells in order to observe the fate of the irradiated spindle and the ability of the cell to continue through mitosis. Cells of the rat kangaroo line (PTK2) were stably transfected by ECFP-tubulin and, using fluorescent microscopy and the automated RoboLase microscope, (Botvinick and Berns, 2005, Micros. Res. Tech. 68:65-74) brightly fluorescent individual cells in metaphase were irradiated with 0.2447 nJ/micropulse corresponding to an irradiance of 1.4496*10^7 J/(ps*cm^2) . Upon irradiation the exposed part of the mitotic spindle immediately lost fluorescence and the following events were observed in the cells over time: (1) immediate contraction of the spindle pole towards the cut, (2) recovery of connection between pole and cut microtubule, (3) completion of mitosis. This system should be very useful in studying internal cellular dynamics of the mitotic spindle.

Recently, we have reported self-rotation of normal red blood cells (RBC), suspended in hypertonic buffer, and trapped in unpolarized laser tweezers. Here, we report use of such an optically driven RBC-motor for microfluidic applications such as pumping/centrifugation of fluids. Since the speed of rotation of the RBC-motor was found to vary with the power of the trapping beam, the flow rate could be controlled by controlling the laser power. In polarized optical tweezers, preferential alignment of trapped RBC was observed. The aligned RBC (simulating a disk) in isotonic buffer, could be rotated in a controlled manner for use as a microfluidic valve by rotation of the plane of polarization of the trapping beam. The thickness of the discotic RBC could be changed by changing the osmolarity of the solution and thus the alignment torque on the RBC due to the polarization of the trapping beam could be varied. Further, in polarized tweezers, the RBCs in hypertonic buffer showed rocking motion while being in rotation. Here, the RBC rotated over a finite angular range, stopped for some time at a particular angle, and then started rotating till it was back to the aligned position and this cycle was found repetitive. This can be attributed to the fact that though the RBCs were found to experience an alignment torque to align with plane of polarization of the tweezers due to its form birefringence, it was smaller in magnitude as compared to the rotational torque due to its structural asymmetry in hypertonic solution. Changes in the laser power caused a transition from/to rocking to/from motor behavior of the RBC in a linearly polarized tweezers. By changing the direction of polarization caused by rotation of an external half wave plate, the stopping angle of rocking could be changed. Further, RBCs suspended in intermediate hypertonic buffer and trapped with polarized tweezers showed fluttering about the vertical plane.

In Selective Laser Micro Sintering the powder particles should be assembled or arranged and sintered together. Optical tweezers make used of optical refractive force to manipulate micro objects. Currently the manipulated objects are limited to nano or several micro meters scale. In this paper we develop a novel optical tweezers which employs pulse laser force to drive bigger particles and assemble them. This pulse laser is controlled to form spiral trap which can grasp big particles. In our experiment the 50μm- 100μm-diameter metal particles were moved on a solid surfaces in a process we call 'laser spiral driving force'. Nearly any shape particle, including sphere and non-regular shape, can be moved on the surfaces.

The emerging field of micro fluidics is challenged with a desire to pump, move and mix minute amounts of fluid. Such micro devices are operated by means of light matter interaction, namely they can be driven through utilizing birefringence and the polarization of the light as well as the reflection and refraction of light. The latter one enables micro motors to be operated in a tangential setup where the rotors are on axis with an optical waveguide. This has the advantage that the complexity of driving such a device in a lab on a chip configuration is reduced by delivering the driving light by means of a waveguide or fiber optics. In this publication we study a micro motor being driven by a fiber optically delivered light beam. We present experimentally and theoretically how light is getting diffracted when in interaction with the rotors of a turning micro motor. By utilizing the two photon signal from a fluorescein dye being excited by a pulsed femtosecond Laser which was used to drive the motor. Additionally the rotation rate is investigated in dependence of the light field parameters.

The photopolymerization in focused laser light is a modern way how to create three-dimensional microstructures with even sub-micron details. We present how this method can be utilized even in very narrow non-diffracting beams. Combination of the self-healing property of these beams and the narrow core of the non-diffracting beam enables the generation of a very uniform fiber of diameter less than 2 micrometers and lengths of several millimeters. Manufacturing conditions and dimensions of the generated fibers are studied.

We demonstrate phase conjugation by means of degenerate four-wave mixing from a colloidal crystal. The nonlinear medium is provided by a periodic spatial refractive index grating created in a colloidal suspension of dielectric microparticles trapped in the intensity distribution of two nearly copropagating interfering laser beams. Phase conjugation is achieved for a probe beam carrying orbital angular momentum as evidenced by the inversion of the topological charge of a phase singularity within the beam. The e ciency and nonlinear parameters of the colloidal crystal as well as its lattice properties are measured and compared to theoretical predictions and previous experimental work.

Kinetics of J-aggregation of thiacarbocyanine (THIATS) has been investigated by measuring time variation of fluorescence spectrum under the solvent evaporation process of THIATS solution. Fluorescence spectrum of the THIATS J-aggregates changes following the increase of concentration, reflecting the formation and ripening process of J-aggregates. These THIATS J-aggregates are trapped and gathered at the focal point by focusing near-infrared (NIR) laser beam into the solution. Two-photon excited fluorescence from the focal spot is concurrently detected with the same trapping laser beam. Fluorescence spectral change is accelerated by focusing NIR laser beam. This result could be attributed that J-aggregates with higher polarizability are preferably formed in the focal spot of NIR laser. Furthermore, we have succeeded in the deposition of J-aggregates on a glass substrate by optical trapping.

An optical conveyor belt created by a standing wave can be used to deliver Brownian particles in a controlled way. The dependence of the particle speed on the speed of traveling standing wave is not simple but two basic modes can be distinguished. For very slow standing wave motion the particle is tightly coupled to the potential, i.e. it "surfs along with the potential wave" and therefore it is called the "Brownian surfer". For bigger standing wave velocities the particle behaves like a swimmer afloat on the surface of the ocean and thus it is called the "Brownian swimmer". The mutual speed of the particle and the standing wave is studied experimentally and theoretically.

In this report, we combine total internal reflection-fluorescence correlation spectroscopy (TIR-FCS) with a single optical trap to simultaneously manipulate and measure the dynamics of individual molecules near the substrate-solution interface. As a proof of principle, polystyrene particles (84 nm in diameter) are used as a model system to test our approach in studying their diffusion properties near surfaces, which are treated with polyethylene glycol 8000, bovine serum albumin or sodium hydroxide. The evanescent field of 543 nm excitation propagates ~100 nm into the solution, and the fluorescence detection is spatially confined by a 25 or 50 μm pinhole that is parfocal with the specimen plane. The optical trap is generated using a cw Ti:sapphire laser at 780 nm. Our results indicate that the particles' diffusion is influenced by surface interactions, which might have further implications on biomembrane studies. Furthermore, the observed translational diffusion of individual particles can be manipulated using an optical trap. By combining the single molecule sensitivity of TIR-FCS with a noninvasive manipulation method, such as optical trapping, we will be able to probe molecular dynamics in biomimetic systems and living cells.

Here, we present real-space studies of Brownian hard sphere transport though externally defined potential energy landscapes. Specifically, we examine how colloidal particles are re-routed as moderately dense suspensions pass through optical lattices, concentrating our attention upon the degree of sorting that occurs in multi-species flows. While methodologies reported elsewhere for microfluidic sorting of colloidal or biological matter employ active intervention to identify and selectively re-route particles one-by-one, the sorting described here is passive, with intrinsically parallel processing. In fact, the densities of co-flowing species examined here are sufficient to allow for significant many-body effects, which generally reduce the efficiencies of re-routing and sorting. We have studied four classes of transport phenomena, involving colloidal traffic within, respectively, a static lattice with a DC fluid flow, a continuously translating lattice with a DC fluid flow, a flashing lattice with AC fluid flow, and a flashing lattice with combined AC and DC fluid flow. We find that continuous lattice translation helps to reduce nearest neighbor particle distances, providing promise for efficiency improvements in future high throughput applications.

At the microscopic scale, the light matter interaction may organize colloidal matter due to a process known as optical binding. Optical binding has now been established as an important issue for the assembly of colloidal matter by light. In the paper we investigate one dimensional optically bound matter of microscopic objects. We develop a dual beam optical fiber trap using a femtosecond laser where the peak power permits us to use two-photon excitation within the host medium. In this trap we can visualize the field distribution in an optical bound array. A numerical model is presented which provides a comparison between theory and experiment.

The paper describes the design of an inexpensive holographic optical tweezers setup. The setup is accompanied by software that allows real-time manipulation of the sample and takes into account the experimental features of the setup, such as aberration correction and LCD modulation. The LCD, a HoloEye LCR-2500, is the physical support of the holograms, which are calculated using the fast random binary mask algorithm. The real-time software achieves 12 fps at full LCD resolution (including aberration correction and modulation) when run on a Pentium IV HT, 3.2 GHz computer.

We report a simple and economical technique to create a wide variety of laser pattern for optical tweezing. The main feature of this technique is a reflective gold mirror that is mounted on a stretched latex membrane which can be vibrated with sound wave at a frequency within 100~600Hz. Due to the vibrating gold mirror, laser beam that is reflected off the mirror exhibits a wide variety of controlled patterns. With the reflected laser pattern directed into an optical microscope for optical tweezing, we were able to form different dynamic configurations of structures made of colloidal microspheres. Different formations of colloidal microspheres that correspond to the reflected laser patterns created by the sound-vibrated mirror have been observed.

A holographic optical tweezers system is constructed using time-shared multiplexing to generate multiple optical traps. Computer generated Fresnel zone plates are used to produce the required laser focuses for standard optical traps while helical zone plates are used to generate Laguerre-Gaussian (LG) laser modes that produce doughnut shaped focuses. Standard Fresnel zone plates are used for trapping non-biological matter whereas LG modes are used in trapping delicate biological matter that is susceptible to damage from excessive laser radiation. A reflective high speed ferroelectric spatial light modulator (SLM), which is used to display the zone plate images, is capable of multiplexing a maximum of 24 binary images at a refresh rate of up to 1440Hz. By programming the SLM to display one binary hologram per bit plane, a commercial 24 bit colour SLM is used to provide 24 multiplexed traps. The ferroelectric holographic system enables multiple independently movable traps using time-shared multiplexing without the need for mechanical movement within the system. The software developed to display the zone plates uses Open GL acceleration to allow fast smooth movement in real time. Open GL utilises the graphics processing unit (GPU) common on many computers today which greatly increases the frame rate of the images displayed on the SLM. Use of the apparatus is demonstrated by the trapping and manipulation of colloidal particles and yeast cells. Preliminary results indicate that the use of LG zone plates for trapping biological cells results in significantly less damage than standard Fresnel zone plates.

We have developed an optical tweezers system by on-demand operation. In this work, we achieved real-time displacement of a trapped particle together with a fixed trapped array of particles. The displacement of a trapping beam spot was performed by introducing the phase shift method. The coexistence of the displacing beam spot with the fixed array of beam spots was realized by time-multiplexing operation of a spatial light modulator.

Liquid crystal (LC) adaptive optical elements are described, which provide an alternative to existing micropositioning technologies in optical tweezing. A full description of this work is given in [1]. An adaptive LC prism supplies tip/tilt to the phase profile of the trapping beam, giving rise to an available steering radius within the x-y plane of 10 μm. Additionally, a modally addressed adaptive LC lens provides defocus, offering a z-focal range for the trapping site of 100 μm. The result is full three-dimensional positional control of trapped particle(s) using a simple and wholly electronic control system. Compared to competing technologies, these devices provide a lower degree of controllability, but have the advantage of simplicity, cost and light efficiency. Furthermore, due to their birefringence, LC elements offer the opportunity of the creation of dual optical traps with controllable depth and separation.

We discuss a new technique to generate force gradient in arrays of optical traps. The arrays can be configured in two or three dimensions by means of phase diffractive optical elements displayed on a spatial light modulator. The design of the diffractive optical elements is based on the approach of spherical wave propagation and superposition, which enables to individually control the strength of each optical trap. Computer simulation and experimental results are discussed for two and three dimensional arrays of traps. An example with silica micro-beads trapped with different forces in two different planes is presented to demonstrate the validity of our approach.

Holographic beam forming to generate and control multiple optical traps has proved successful using high-resolution spatial light modulators (SLMs). This type of beam control allows a multitude of traps to be independently controlled in three dimensions. Also, exotic beam shapes and profiles can be generated, which gives the optical trapping system even greater flexibility. Until recently, the generation of high resolution phase patterns has limited the speed of dynamic holographic optical trapping (HOT) systems. Today, video rate operation controlling hundreds of traps using 512x512 phase masks is possible and significantly faster operation is possible with fewer traps using less phase resolution. Therefore, phase-only liquid crystal modulator response is becoming the bottleneck. This paper discusses recent advances in SLM developments which address this issue.

Digital holography enables the creation of multiple optical traps at arbitrary three-dimensional locations and spatial light modulators permit updating those holograms at video rates. However, the time required for computing the holograms makes interactive optical manipulation of several samples difficult to achieve. We introduce an algorithm for computing holographic optical tweezers that is both easy to implement and capable of speeds in excess of 10 Hz when running on a Pentium IV computer. A discussion of the pros and cons of the algorithm, a mathematical analysis of the efficiency of the resulting traps, as well as results of the three-dimensional manipulation of polystyrene micro spheres are included.

Holographic or diffractive optical components, such as a spatial light modulator (SLM), can be used in optical tweezers for the creation of multiple and modified optical traps. In addition to this, SLMs can also be used to correct for aberrations within the optical train resulting in an improved trapping performance. Typically an electrically addressed SLM may deviate from flatness by up to 4λ, dominated by astigmatism due to the overall curvature of the SLM surface. This astigmatism may be corrected by adding the appropriate hologram to the SLM display resulting in a dramatic improvement in the fidelity of the focussed spot. The impact that this correction has on the performance of the optical trap is most noticeable for small particles. For the SLM used in this study, the improvement in trap performance for a 0.8 μm diameter particles can be in excess of 25%. However, for 5 μm diameter particles our results show an improvement of less than 0.5%. This dependence upon particle size is most probably associated with the relative size of the PSF and the trapped particle. Once the PSF is significantly smaller than the particle diameter, further reduction brings little improvement in trap performance.

The three-dimensional forces acting on a dielectric microparticle illuminated by two counterpropagating beams with variable intensity profiles are theoretically studied. Size-adjustable intensity profiles of the constituent beams are easily implemented using the generalized phase contrast (GPC) method. Our numerical calculations include the dependence of the axial and transverse trapping forces with location of the microsphere between the opposing beams. These numerical results show potential improvement in the large dynamic range for axial position control of microparticles in GPC-based counterpropagating-beam traps - achieved with the use of size-varied constituent tophat beams. We also assess the quality of the obtained trap potentials by calculating the axial and transverse stiffness associated with the optical traps.

We demonstrate laser manipulation of different nematic and smectic liquid crystal droplets in heavy water. Peculiarities of laser trapping and manipulation depending on the molecular structure of liquid crystal are discussed. Possibility of molecular reordering inside the tweezed droplet is demonstrated. This phenomenon can be used to induce the birefringence and to manipulate droplets, which have a small birefringence, e.g., a radial droplet of negligible birefringence can be turned into a birefringent one at high laser trapping power. This is a demonstration of the optical nonlinear effect being responsible for a micro-mechanical phenomenon such as spinning of the droplet in a circularly polarized laser tweezers. Three-photon absorption of MBAPB dye at 1064 nm wavelength is demonstrated inside the liquid crystal droplet at comparatively low ~60 MW/cm² intensity.