Side-view holographic endomicroscopy via a custom-terminated multimode fibre.

Microendoscopes based on optical fibres have recently come to the fore as promising candidates allowing in-vivo observations of otherwise inaccessible biological structures in animal models. Despite being still in its infancy, imaging can now be performed at the tip of a single multimode fibre, by relying on powerful holographic methods for light control. Fibre based endoscopy is commonly performed en face, resulting in possible damage of the specimen owing to the direct contact between the distal end of the probe and target. On this ground, we designed an all-fibre probe with an engineered termination that reduces compression and damage to the tissue under investigation upon probe insertion. The geometry of the termination brings the field of view to a plane parallel to the fibre's longitudinal direction, conveying the probe with off-axis imaging capabilities. We show that its focusing ability also benefits from a higher numerical aperture, resulting in imaging with increased spatial resolution. The effect of probe insertion was investigated inside a tissue phantom comprising fluorescent particles suspended in agarose gel, and a comparison was established between the novel side-view probe and the standard en face fibre probe. This new concept paves the way to significantly less invasive deep-tissue imaging.

Computational image enhancement of multimode fibre-based holographic endo-microscopy: harnessing the muddy modes.

In imaging geometries, which employ wavefront-shaping to control the light transport through a multi-mode optical fibre (MMF), this terminal hair-thin optical component acts as a minimally invasive objective lens, enabling high resolution laser-scanning fluorescence microscopy inside living tissues at depths hardly accessible by any other light-based technique. Even in the most advanced systems, the diffraction-limited foci scanning the object across the focal plane are contaminated by a stray optical signal carrying typically few tens of % of the total optical power. The stray illumination takes the shape of a randomised but reproducible speckle, and is unique for each position of the focus. We experimentally demonstrate that the performance of imaging a fluorescent object can be significantly improved, when resulting images are computationally post-processed, utilising records of intensities of all speckle-contaminated foci used in the imaging procedure. We present two algorithms based on a regularised iterative inversion and regularised direct pseudo-inversion respectively which lead to enhancement of the image contrast and resolution.

Diffusing up the Hill: Dynamics and Equipartition in Highly Unstable Systems.

Stochastic motion of particles in a highly unstable potential generates a number of diverging trajectories leading to undefined statistical moments of the particle position. This makes experiments challenging and breaks down a standard statistical analysis of unstable mechanical processes and their applications. A newly proposed approach takes advantage of the local characteristics of the most probable particle motion instead of the divergent averages. We experimentally verify its theoretical predictions for a Brownian particle moving near an inflection in a highly unstable cubic optical potential. The most likely position of the particle atypically shifts against the force, despite the trajectories diverging in the opposite direction. The local uncertainty around the most likely position saturates even for strong diffusion and enables well-resolved position detection. Remarkably, the measured particle distribution quickly converges to a quasistationary one with the same atypical shift for different initial particle positions. The demonstrated experimental confirmation of the theoretical predictions approves the utility of local characteristics for highly unstable systems which can be exploited in thermodynamic processes to uncover energetics of unstable systems.

Omnidirectional Transport in Fully Reconfigurable Two Dimensional Optical Ratchets.

A fully reconfigurable two-dimensional (2D) rocking ratchet system created with holographic optical micromanipulation is presented. We can generate optical potentials with the geometry of any Bravais lattice in 2D and introduce a spatial asymmetry with arbitrary orientation. Nontrivial directed transport of Brownian particles along different directions is demonstrated numerically and experimentally, including on axis, perpendicular, and oblique with respect to an unbiased ac driving. The most important aspect to define the current direction is shown to be the asymmetry and not the driving orientation, and yet we show a system in which the asymmetry orientation of each potential well does not coincide with the transport direction, suggesting an additional symmetry breaking as a result of a coupling with the lattice configuration. Our experimental device, due to its versatility, opens up a new range of possibilities in the study of nonequilibrium dynamics at the microscopic level.

Direct measurement of the temperature profile close to an optically trapped absorbing particle.

The surface temperature of an absorbing particle trapped in optical tweezers (OTs) is measured using a mixture of two fluorescent dyes. We analyze the dependence of temperature on both laser power and the radial distance from its surface, and we verify the 1/r decrease of temperature with increasing distance from the particle surface. We detect the variations of spectral profiles as the medium temperature changes. The temperature dependent signal, i.e., the ratio of summed intensities from two distinct spectral regions, is affected by the convolution of temperature profile with transfer function of the spectroscopic system. We analyze this effect and determine the temperature increase on the surface of a core-shell particle trapped by OTs.

Complex rotational dynamics of multiple spheroidal particles in a circularly polarized, dual beam trap.

We examine the rotational dynamics of spheroidal particles in an optical trap comprising counter-propagating Gaussian beams of opposing helicity. Isolated spheroids undergo continuous rotation with frequencies determined by their size and aspect ratio, whilst pairs of spheroids display phase locking behaviour. The introduction of additional particles leads to yet more complex behaviour. Experimental results are supported by numerical calculations.

Rotation, oscillation and hydrodynamic synchronization of optically trapped oblate spheroidal microparticles.

While the behavior of optically trapped dielectric spherical particles has been extensively studied, the behavior of non-spherical particles remains mainly unexplored. In this work we focus on the dynamics of oblate spheroidal particles trapped in a tightly focused elliptically-polarized vortex beam. In our experiments we used polystyrene spheroids of aspect ratio of major to minor axes equal to 2.55 and of a volume equal to a sphere of diameter 1.7µm. We demonstrate that such particles can be trapped in three dimensions, with the minor axis oriented perpendicular to both the beam polarization (linear) and the beam propagation, can spin in a circularly polarized beam and an optical vortex beam around the axis parallel with the beam propagation. We also observed that these particles can exhibit a periodic motion in the plane transversal to the beam propagation. We measured that the transfer of the orbital angular momentum from the vortex beam to the spheroid gives rise to torques one order of magnitude stronger comparing to the circularly polarized Gaussian beam. We employed a phase-only spatial light modulator to generate several vortex beam traps with one spheroid in each of them. Due to independent setting of beams parameters we controlled spheroids frequency and sense of rotation and observed hydrodynamic phase and frequency locking of rotating spheroids. These optically driven spheroids offer a simple alternative approach to the former techniques based on birefringent, absorbing or chiral microrotors.

Optical sorting of nonspherical and living microobjects in moving interference structures.

Contactless, sterile and nondestructive separation of microobjects or living cells is demanded in many areas of biology and analytical chemistry, as well as in physics or engineering. Here we demonstrate advanced sorting methods based on the optical forces exerted by travelling interference fringes with tunable periodicity controlled by a spatial light modulator. Besides the sorting of spherical particles we also demonstrate separation of algal cells of different sizes and particles of different shapes. The three presented methods offer simultaneous sorting of more objects in static suspension placed in a Petri dish or on a microscope slide.

Optical manipulation of aerosol droplets using a holographic dual and single beam trap.

We present optical trapping and manipulation of pure water and salt water airborne droplets of various sizes ranging from sub-micrometers up to several tens of micrometers in a holographic dual and single beam trap. In the dual beam trap, successful fusion of droplets as well as precise delivery of many droplets and manipulation of multiple droplets are demonstrated. Furthermore, employing the transfer of the orbital angular momentum of light from Laguerre-Gaussian beams, we show that the water droplets orbit around the beam propagation axis and their tangential speed can be controlled by beam waist magnitude. We also demonstrate that sub-micrometer sized pure water droplets can be trapped and manipulated by a single beam trap with a relatively low numerical aperture. In this case, multiple stable trapping positions were observed, both theoretically and experimentally, which were due to the optical intensity oscillations in the focal region of the laser beam.

Following the mechanisms of bacteriostatic versus bactericidal action using Raman spectroscopy.

Antibiotics cure infections by influencing bacterial growth or viability. Antibiotics can be divided to two groups on the basis of their effect on microbial cells through two main mechanisms, which are either bactericidal or bacteriostatic. Bactericidal antibiotics kill the bacteria and bacteriostatic antibiotics suppress the growth of bacteria (keep them in the stationary phase of growth). One of many factors to predict a favorable clinical outcome of the potential action of antimicrobial chemicals may be provided using in vitro bactericidal/bacteriostatic data (e.g., minimum inhibitory concentrations-MICs). Consequently, MICs are used in clinical situations mainly to confirm resistance, and to determine the in vitro activities of new antimicrobials. We report on the combination of data obtained from MICs with information on microorganisms' "fingerprint" (e.g., DNA/RNA, and proteins) provided by Raman spectroscopy. Thus, we could follow mechanisms of the bacteriostatic versus bactericidal action simply by detecting the Raman bands corresponding to DNA. The Raman spectra of Staphylococcus epidermidis treated with clindamycin (a bacteriostatic agent) indeed show little effect on DNA which is in contrast with the action of ciprofloxacin (a bactericidal agent), where the Raman spectra show a decrease in strength of the signal assigned to DNA, suggesting DNA fragmentation.

Synchronization of spin-driven limit cycle oscillators optically levitated in vacuum.

We explore, experimentally and theoretically, the emergence of coherent coupled oscillations and synchronization between a pair of non-Hermitian, stochastic, opto-mechanical oscillators, levitated in vacuum. Each oscillator consists of a polystyrene microsphere trapped in a circularly polarized, counter-propagating Gaussian laser beam. Non-conservative, azimuthal forces, deriving from inhomogeneous optical spin, push the micro-particles out of thermodynamic equilibrium. For modest optical powers each particle shows a tendency towards orbital circulation. Initially, their stochastic motion is weakly correlated. As the power is increased, the tendency towards orbital circulation strengthens and the motion of the particles becomes highly correlated. Eventually, centripetal forces overcome optical gradient forces and the oscillators undergo a collective Hopf bifurcation. For laser powers exceeding this threshold, a pair of limit cycles appear, which synchronize due to weak optical and hydrodynamic interactions. In principle, arrays of such Non-Hermitian elements can be arranged, paving the way for opto-mechanical topological materials or, possibly, classical time crystals. In addition, the preparation of synchronized states in levitated optomechanics could lead to new and robust sensors or alternative routes to the entanglement of macroscopic objects.

110 µm thin endo-microscope for deep-brain in vivo observations of neuronal connectivity, activity and blood flow dynamics.

Light-based in-vivo brain imaging relies on light transport over large distances of highly scattering tissues. Scattering gradually reduces imaging contrast and resolution, making it difficult to reach structures at greater depths even with the use of multiphoton techniques. To reach deeper, minimally invasive endo-microscopy techniques have been established. These most commonly exploit graded-index rod lenses and enable a variety of modalities in head-fixed and freely moving animals. A recently proposed alternative is the use of holographic control of light transport through multimode optical fibres promising much less traumatic application and superior imaging performance. We present a 110 µm thin laser-scanning endo-microscope based on this prospect, enabling in-vivo volumetric imaging throughout the whole depth of the mouse brain. The instrument is equipped with multi-wavelength detection and three-dimensional random access options, and it performs at lateral resolution below 1 µm. We showcase various modes of its application through the observations of fluorescently labelled neurones, their processes and blood vessels. Finally, we demonstrate how to exploit the instrument to monitor calcium signalling of neurones and to measure blood flow velocity in individual vessels at high speeds.

Optical forces induced behavior of a particle in a non-diffracting vortex beam.

An interaction between a light field with complex field spatial distribution and a micro-particle leads to forces that drag the particle in space and may confine it in a stable position or a trajectory. The particle behavior is determined by its size with respect to the characteristic length of the spatially periodic or symmetric light field distribution. We study theoretically and experimentally the behavior of a microparticle near the center of an optical vortex beam in a plane perpendicular to the beam propagation. We show that such particle may be stably trapped either in a dark spot on the vortex beam axis, or in one of two points placed off the optical axis. It may also circulate along a trajectory having its radius smaller or equal to the radius of the first bright vortex ring.

Spin to orbital light momentum conversion visualized by particle trajectory.

In a tightly focused beam of light having both spin and orbital angular momentum, the beam exhibits the spin-orbit interaction phenomenon. We demonstrate here that this interaction gives rise to series of subtle, but observable, effects on the dynamics of a dielectric microsphere trapped in such a beam. In our setup, we control the strength of spin-orbit interaction with the width, polarization and vorticity of the beam and record how these parameters influence radius and orbiting frequency of the same single orbiting particle pushed by the laser beam. Using Richard and Wolf model of the non-paraxial beam focusing, we found a very good agreement between the experimental results and the theoretical model based on calculation of the optical forces using the generalized Lorenz-Mie theory extended to a non-paraxial vortex beam. Especially the radius of the particle orbit seems to be a promising parameter characterizing the spin to orbital momentum conversion independently on the trapping beam power.

Thermally induced micro-motion by inflection in optical potential.

Recent technological progress in a precise control of optically trapped objects allows much broader ventures to unexplored territory of thermal motion in non-linear potentials. In this work, we exploit an experimental set-up of holographic optical tweezers to experimentally investigate Brownian motion of a micro-particle near the inflection point of the cubic optical potential. We present two complementary views on the non-linear Brownian motion. On an ensemble of stochastic trajectories, we simultaneously determine (i) the detailed short-time position statistics and (ii) the long-distance first-passage time statistics. We evaluate specific statistical moment ratios demonstrating strongly non-linear stochastic dynamics. This is a crucial step towards a possible massive exploitation of the broad class of complex non-linear stochastic effects with objects of more complex structure and shape including living ones.