[Show abstract][Hide abstract] ABSTRACT: It is shown that grisms, a grating and prism combination, are a simple way to achieve chromatic control in 3D multi-plane imaging. A pair of grisms, whose separation can be varied, provide a collimated beam with a tuneable chromatic shear from a collimated polychromatic input. This simple control permits the correction of chromatic smearing in 3D imaging using off-axis Fresnel zone plates and improved control of the axial profile of a focussed spot in multi-photon experiments.
[Show abstract][Hide abstract] ABSTRACT: We extend the redundant spacings calibration method for finding piston coefficients affecting the elements of a dilute aperture array so that tilt phase coefficients can also be calculated and corrected without the need for assumptions about the object. The tilt coefficient retrieval method is successfully demonstrated in simulation, and the specifics of correction by image sharpness are discussed, showing that in dilute aperture systems this method does not necessarily produce a unique image.
Journal of the Optical Society of America A 05/2012; 29(5):757-66. · 1.67 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Image denoising and signal enhancement are two common steps to improve particle contrast for detection in low-signal-to-noise ratio (SNR) fluorescence live-cell images. However, denoising may oversmooth features of interest, particularly weak features, leading to false negative detection. Here, we propose a robust framework for particle detection in which image denoising in the grayscale image is not needed, so avoiding image oversmoothing. A key to our approach is the new development of a particle enhancement filter based on the recently proposed particle probability image to obtain significantly enhanced particle features and greatly suppressed background in low-SNR and low-contrast environments. The new detection method is formed by combining foreground and background markers with watershed transform operating in both particle probability and grayscale spaces; dynamical switchings between the two spaces can optimally make use the information in images for accurate determination of particle position, size, and intensity. We further develop the interacting multiple mode filter for particle motion modeling and data association by incorporating the extra information obtained from our particle detector to enhance the efficiency of multiple particle tracking. We find that our methods lead to significant improvements in particle detection and tracking efficiency in fluorescence live-cell applications.
[Show abstract][Hide abstract] ABSTRACT: We describe a method for tracking the position of small features in three dimensions from images recorded on a standard microscope with an inexpensive attachment between the microscope and the camera. The depth-measurement accuracy of this method is tested experimentally on a wide-field, inverted microscope and is shown to give approximately 8 nm depth resolution, over a specimen depth of approximately 6 µm, when using a 12-bit charge-coupled device (CCD) camera and very bright but unresolved particles. To assess low-flux limitations a theoretical model is used to derive an analytical expression for the minimum variance bound. The approximations used in the analytical treatment are tested using numerical simulations. It is concluded that approximately 14 nm depth resolution is achievable with flux levels available when tracking fluorescent sources in three dimensions in live-cell biology and that the method is suitable for three-dimensional photo-activated localization microscopy resolution. Sub-nanometre resolution could be achieved with photon-counting techniques at high flux levels.
Journal of The Royal Society Interface 01/2011; 8(60):942-51. · 4.91 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: This paper discusses a range of phase-diversity and tracking applications that have been demonstrated experimentally, and will present an analysis of experimental errors associated with the system used. Simultaneous imaging in multiple imaging modes is demonstrated and the use of wavefront-sensing techniques to achieve nanometric depth resolution is reviewed.
[Show abstract][Hide abstract] ABSTRACT: Fluorescence imaging of dynamical processes in live cells often results in a low signal-to-noise ratio. We present a novel feature-preserving non-local means approach to denoise such images to improve feature recovery and particle detection. The commonly used non-local means filter is not optimal for noisy biological images containing small features of interest because image noise prevents accurate determination of the correct coefficients for averaging, leading to over-smoothing and other artifacts. Our adaptive method addresses this problem by constructing a particle feature probability image, which is based on Haar-like feature extraction. The particle probability image is then used to improve the estimation of the correct coefficients for averaging. We show that this filter achieves higher peak signal-to-noise ratio in denoised images and has a greater capability in identifying weak particles when applied to synthetic data. We have applied this approach to live-cell images resulting in enhanced detection of end-binding-protein 1 foci on dynamically extending microtubules in photo-sensitive Drosophila tissues. We show that our feature-preserving non-local means filter can reduce the threshold of imaging conditions required to obtain meaningful data.
Journal of Structural Biology 12/2010; 172(3):233-43. · 3.36 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: A conventional microscope produces a sharp image from just a single object-plane. This is often a limitation, notably in cell biology. We present a microscope attachment which records sharp images from several object-planes simultaneously. The key concept is to introduce a distorted diffraction grating into the optical system, establishing an order-dependent focussing power in order to generate several images, each arising from a different object-plane. We exploit this multiplane imaging not just for bio-imaging but also for nano-particle tracking, achieving approximately 10 nm z position resolution by parameterising the images with an image sharpness metric.
[Show abstract][Hide abstract] ABSTRACT: The principle of redundant spacings calibration has previously been described for the purpose of calibrating piston phase aberration affecting elements of a dilute aperture array using a system of linear equations in terms of the aperture phases as well as object phase information. Here we develop matrices for the correction of piston phase aberration by use of image sharpness and also by phase retrieval. These are both presented in wavefront sensor formulation in order to draw analogy between the approaches. We then discuss solution ambiguity affecting both methods and describe array design criteria to prevent such ambiguity. The problem of increased image aliasing under image sharpness correction is also highlighted.
Journal of the Optical Society of America A 02/2009; 26(1):195-205. · 1.67 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: We present the application of wavefront sensing to three-component, three-dimensional micro particle tracking velocimetry
(μPTV). The technique is based upon examining the defocus of the wavefront scattered by a tracer particle and from such information
establishing the 3-D tracer location. The imaging system incorporates a cylindrical lens acting as an anamorphic element that
creates different magnifications in the two orthogonal axes. A single anamorphic image is obtained from each tracer, which
contains sufficient information to reconstruct the wavefront defocus and uniquely identify the tracer’s axial position. A
mathematical model of the optical system is developed and shows that the lateral and depth performance of the sensor can be
largely independently varied across a wide range. Hence, 3-D image resolution can be achieved from a single viewpoint, using
simple and inexpensive optics and applied to a wide variety of microfluidic or biological systems. Our initial results show
that an uncertainty in depth of 0.18 μm was achieved over a 20-μm range. The technique was employed to measure the 3-D velocity
field of micron-sized fluorescent tracers in a flow within a micro channel, and an uncertainty of 2.8 μm was obtained in the
axial direction over a range of 500 μm. The experimental results were in agreement with the expected fluid flow when compared
to the corresponding CFD model. Thus, wavefront sensing proved to be an effective approach to obtain quantitative measurements
of three-component three-dimensional flows in microfluidic devices.
Experiments in Fluids 01/2009; 47(4-5):849-863. · 1.57 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Pupil replication and hypertelescope systems for imaging telluric exoplanets in scattered light are treated. Analytic expressions for the spread functions in one and two dimensions and in the presence of various forms of error are given. Error effects considered include aperture misalignment, tilts, piston, pointing errors, and unequal beam amplitude. The performance of the two approaches is contrasted, and the analytic results are compared with simulation results.
The Astrophysical Journal 12/2008; 658(2):1380. · 6.73 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Pupil replication is a new optical technique that decreases the diameter of the image of a star on the optical axis. With pupil replication, one can improve the suppression of starlight in high dynamic range exoplanet imaging and image planets closer to their host star. Pupil replication is an auxiliary technique intended to improve the imaging spectroscopic performance of coronographic or pupil apodization techniques. Here we introduce pupil replication and use numerical simulations to show that it may be combined with pupil apodization to improve exoplanet detection and spectroscopy.
The Astrophysical Journal 12/2008; 618(2):L165. · 6.73 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: We use a two-dimensional deformable mirror to shape the spatial profile of an ultrafast laser beam that is then used to inscribe structures in a soda-lime silica glass slide. By doing so we demonstrate that it is possible to control the asymmetry of the cross section of ultrafast laser inscribed optical waveguides via the curvature of the deformable mirror. When tested using 1.55 mum light, the optimum waveguide exhibited coupling losses of approximately 0.2 dB/facet to Corning SMF-28 single mode fiber and propagation losses of approximately 1.5 dB.cm(-1). This technique promises the possibility of combining rapid processing speeds with the ability to vary the waveguide cross section along its length.
[Show abstract][Hide abstract] ABSTRACT: We propose an annular-aperture-based defocusing technique for three-dimensional (3D) particle metrology from a single camera view. This simple configuration has high optical efficiency and the ability to deal with overlapped defocused images. Initial results show that an uncertainty in depth of 23 microm can be achieved over a range of 10 mm for macroscopic systems. This method can also be applied in microscopy for the measurement of fluorescently doped microparticles, thus providing a promising solution for 3D flow metrology at both macroscales and microscales.
[Show abstract][Hide abstract] ABSTRACT: Observing biological processes in real-time and in single live cells is a vital step towards understanding cell behaviour and the way cells interact with the world around them, However, this requires real time three dimensional (4D) tracking of nanoparticles which is challenging and traditionally relies on sequential capture of 2D images to construct a 3D picture. We discuss a new approach to 4D nanoparticle tracking that utilises a specially designed diffraction grating which behaves as a lens with a different focal length in each diffraction order thereby producing pseudo D imaging over the imaged field. The current experimental system has the ability to track a single particle in a 50×50×6mum volume, with an accuracy of better than 50 nm in each dimension.
[Show abstract][Hide abstract] ABSTRACT: Thin film metrology is an important quality control mechanism used in many industrial processes. Conventional techniques of Ellipsometry and Spectral Reflectance are limited in applicability to in situ measurements during film manufacture. This paper considers the measurement of wavefront shape reflected from interfaces within a laminate for determination of layer thickness and profile. For thick layers a wavefront sensor can provide the requisite wavefront shape data, for thin layers the interference between reflected wavefronts is used to retrieve wavefront shape.
[Show abstract][Hide abstract] ABSTRACT: We introduce an approach based on using an iterative simulated annealing algorithm to drive the outcome of femtosecond laser ablation towards a specific target shape by using a two-dimensional deformable mirror. Unlike previous work combining adaptive optics and laser machining we use the machining outcome itself as the fitness parameter for the optimization procedure. Single-pulse ablation features with programmable aspect ratios and dimensions as small as 2.5 µm are faithfully reproduced by the technique in a chromium-on-glass test sample.
Journal of Optics A Pure and Applied Optics 11/2007; 9:1100-1104. · 1.92 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: We study an adaptive dynamical feedback system for phasing of segmented mirrors. The system, which combines a Zernike wave front sensor with a feedback loop, is shown to be capable of separating (static) misalignment errors of segmented mirrors from (dynamical) atmospheric turbulence. Our numerical simulations establish that it can retrieve multi-wave segment misalignment error in the presence of large-amplitude atmospheric phase distortion.
Applied Physics B 12/2006; 86(1):139-145. · 1.78 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: We present the application of wavefront sensing to particle image velocimetry for three-component (3C), three-dimensional (3D) flow measurement from a single view. The technique is based upon measuring the wavefront scattered by a tracer particle and from that wavefront the 3D tracer location can be determined. Hence, from a temporally resolved sequence of 3D particle locations the velocity vector field is obtained. Two approaches to capture the data required to measure the wavefronts are described: multi-planar imaging using a distorted diffraction grating and an anamorphic technique. Both techniques are optically efficient, robust and compatible with coherent and incoherent scattering from flow tracers. The depth (range) resolution and repeatability have been quantified experimentally using a single mode fiber source representing a tracer particle. The anamorphic approach is shown to have the greatest measurement range and hence was selected for the first proof of principle experiments using this technique for 3D particle imaging velocimetry (PIV) on a sparsely seeded gas phase flow.
Experiments in Fluids 12/2006; 41(6):881-891. · 1.57 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The design, testing and operation of a system for telecentric 3-dimensional imaging of dynamic objects is presented. The simple system is capable of rapid electronic scanning of a single focal plane within a specimen or of simultaneous focusing on multiple planes whose depth and relative spacing within the specimen can be changed electronically. Application to studies of dynamic processes in microscopy is considered.