Full-field Microscopy Research Articles

Fast X-ray imaging at beamline I13L at Diamond Light Source

The imaging branch of the dual-branch beamline I13L at Diamond Light Source has been operational since April 2012. This branch is dedicated to hard X-ray imaging (in-line phase contrast radiography and tomography, and full-field microscopy), with energies in the ranges 6-30keV. At present we aim to achieve spatial resolution of the order of 1 μm over a field of view of l-20mm2. This branch aims to excel at imaging experiment of fast dynamic processes, where it is of interest to have short exposure times and high frame rates. To accommodate for this, we prepared for the beamline to operate with "pink" beam to provide higher flux, an efficient detection system, and rapid data acquisition, transfer, and saving to storage. This contributed paper describes the present situation and illustrate the author's goal for the mid-future.

Resonant amplification of vortex-core oscillations by coherent magnetic-field pulses

Vortex structures in soft magnetic nanodisks are highly attractive due to their scientific beauty and potential technological applications. Here, we experimentally demonstrated the resonant amplification of vortex oscillations by application of simple coherent field pulses tuned to optimal width and time intervals. In order to investigate vortex excitations on the sub-ns time scale, we employed state-of-the-art time-resolved full-field soft X-ray microscopy of 70 ps temporal and 25 nm lateral resolution. We found that, due to the resonant enhancement of the vortex gyration motion, the signal input power can be significantly reduced to ~ 1 Oe in field strength, while increasing signal gains, by increasing the number of the optimal field pulses. We identified the origin of this behavior as the forced resonant amplification of vortex gyration. This work represents an important milestone towards the potential implementation of vortex oscillations in future magnetic vortex devices.

Full-field optical coherence microscopy with Riesz transform-based demodulation for dynamic imaging

We present dynamic full-field optical coherence microscope imaging using a scientific complementary metal oxide semiconductor camera in conjunction with a demodulation scheme based on the Riesz transform and monogenic signals. The potential of our approach is verified by a comparison with conventional phase-stepping as well as with an analytic reconstruction method and finally exemplified for dynamic mechanical testing of a polymer/fiber composite structure.

Full-field Hilbert phase microscopy using nearly common-path low coherence off-axis interferometry for quantitative imaging of biological cells

We demonstrate single shot low coherence quantitative Hilbert phase microscopy (HPM) for the reconstruction of the two dimensional (2D) phase map of biological cells. The system is based on a compact and nearly common-path high magnification Mirau-interferometric objective lens. The spatial carrier frequency of the interference fringes was increased by means of introducing tilt in one of the arms of the interferometer, thus making the system off-axis. The system is user friendly as the interference fringes and imaging of objects with high lateral and axial resolution can be obtained quickly using a low cost commercially available microscope. Experimental results for the 2D phase map of polystyrene spheres and human red blood cells (RBCs) are presented. Hilbert transform fringe analysis was used for reconstructing the phase map and refractive index (RI) of the objects. For dynamic substances which change rapidly, single shot low coherence interferometric microscopy is an important method for obtaining the phase. Experimental results with increased field-of-view and large tilt angle are also presented. It is well known that on increasing the tilt angle for improved spatial phase sampling the object remains focused in only a small area even though the field-of-view is large. This limitation was overcome by means of vertical scanning low coherence interferometry. Due to the low coherence properties of the light source the interference occurs only at the desired location of the object, i.e., where the object is sharply focused. The object was vertically scanned and the single shot interferograms were recorded for every scan and analyzed by Hilbert transform. In this way a large area of the sample can be imaged quantitatively.

Optimized Cross-Slot Flow Geometry for Microfluidic Extensional Rheometry

A precision-machined cross-slot flow geometry with a shape that has been optimized by numerical simulation of the fluid kinematics is fabricated and used to measure the extensional viscosity of a dilute polymer solution. Full-field birefringence microscopy is used to monitor the evolution and growth of macromolecular anisotropy along the stagnation point streamline, and we observe the formation of a strong and uniform birefringent strand when the dimensionless flow strength exceeds a critical Weissenberg number Wi(crit) ≈ 0.5. Birefringence and bulk pressure drop measurements provide self-consistent estimates of the planar extensional viscosity of the fluid over a wide range of deformation rates (26 s(-1) ≤ ε ≤ 435 s(-1)) and are also in close agreement with numerical simulations performed by using a finitely extensible nonlinear elastic dumbbell model.

Label-free characterization of living human induced pluripotent stem cells by subcellular topographic imaging technique using full-field quantitative phase microscopy coupled with interference reflection microscopy

There is a need for a noninvasive technique to monitor living pluripotent stem cell condition without any labeling. We present an optical imaging technique that is able to capture information about optical path difference through the cell and cell adhesion properties simultaneously using a combination of quantitative phase microscopy (QPM) and interference reflection microscopy (IRM) techniques. As a novel application of QPM and IRM, this multimodal imaging technique demonstrated its ability to distinguish the undifferentiated status of human induced pluripotent stem (hiPS) cells quantitatively based on the variation of optical path difference between the nucleus and cytoplasm as well as hiPS cell-specific cell adhesion properties.

Zone-doubled Fresnel zone plates for high-resolution hard X-ray full-field transmission microscopy.

Full-field transmission X-ray microscopy is a unique non-destructive technique for three-dimensional imaging of specimens at the nanometer scale. Here, the use of zone-doubled Fresnel zone plates to achieve a spatial resolution better than 20 nm in the hard X-ray regime (8-10 keV) is reported. By obtaining a tomographic reconstruction of a Ni/YSZ solid-oxide fuel cell, the feasibility of performing three-dimensional imaging of scientifically relevant samples using such high-spatial-resolution Fresnel zone plates is demonstrated.

Advantages of a synchrotron bending magnet as the sample illuminator for a wide-field X-ray microscope

In this paper the choice between bending magnets and insertion devices as sample illuminators for a hard X-ray full-field microscope is investigated. An optimized bending-magnet beamline design is presented. Its imaging speed is very competitive with the performance of similar microscopes installed currently at insertion-device beamlines. The fact that imaging X-ray microscopes can accept a large phase space makes them very well suited to the output characteristics of bending magnets which are often a plentiful and paid-for resource. There exist opportunities at all synchrotron light sources to take advantage of this finding to build bending-magnet beamlines that are dedicated to transmission X-ray microscope facilities. It is expected that demand for such facilities will increase as three-dimensional tomography becomes routine and advanced techniques such as mosaic tomography and XANES tomography (taking three-dimensional tomograms at different energies to highlight elemental and chemical differences) become more widespread.

Sequential single-shot imaging of nanoscale dynamic interactions with a table-top soft x-ray laser

We demonstrate the first real-space recording of nanoscale dynamic interactions using single-shot soft x-ray (SXR) full-field laser microscopy. A sequence of real-space flash images acquired with a table-top SXR laser was used to capture the motion of a rapidly oscillating magnetic nanoprobe. Changes of 30 nm in the oscillation amplitude were detected when the nanoprobe was made to interact with stray fields from a magnetic sample. The table-top visualization of nanoscale dynamics in real space can significantly contribute to the understanding of nanoscale processes and can accelerate the development of new nanodevices.

Numerical correction of coherence gate in full-field swept-source interference microscopy

A big problem in low-coherence interference microscopy is the degradation of the coherence signal caused by shift of the angular and temporal spectrum gates. It limits the depth of field in confocal optical coherence microscopy and degrades images of sample inner structure in most interference microscopy techniques. To overcome this problem we propose numerical correction of the coherence gate in application to full-field swept-source interference microscopy. The proposed technique allows three-dimensional sample imaging without mechanical movement of the microscope components and is also capable of determining separately the geometrical thickness and the refractive index of the sample layers, when the sample contains a transversal pattern. The applicability of the proposed technique is verified with numerical simulation.

Full-Field Near-Field Optical Microscope for Cell Imaging

We report a new full-field fluorescence microscopy method for imaging live cell membranes based on supercritical near-field emission. This technique consists of extracting the self-interference between undercritical and supercritical light by simple image subtraction. In the objective back focal plane, this is equivalent to adding a virtual mask blocking the subcritical emission. We show that this virtual mask is radically different from a real physical mask, enabling a 100 nm axial confinement and enhancing the image sensitivity without damaging the lateral resolution. This technique is easy to implement and simultaneously provides images of the inner parts of the cell and its membrane with standard-illumination light.

Logic Operations Based on Magnetic-Vortex-State Networks

Logic operations based on coupled magnetic vortices were experimentally demonstrated. We utilized a simple chain structure consisting of three physically separated but dipolar-coupled vortex-state Permalloy disks as well as two electrodes for application of the logical inputs. We directly monitored the vortex gyrations in the middle disk, as the logical output, by time-resolved full-field soft X-ray microscopy measurements. By manipulating the relative polarization configurations of both end disks, two different logic operations are programmable: the XOR operation for the parallel polarization and the OR operation for the antiparallel polarization. This work paves the way for new-type programmable logic gates based on the coupled vortex-gyration dynamics achievable in vortex-state networks. The advantages are as follows: a low-power input signal by means of resonant vortex excitation, low-energy dissipation during signal transportation by selection of low-damping materials, and a simple patterned-array structure.

Speckle-free laser imaging using random laser illumination

Many imaging applications require increasingly bright illumination sources, motivating the replacement of conventional thermal light sources with light emitting diodes (LEDs), superluminescent diodes (SLDs) and lasers. Despite their brightness, lasers and SLDs are poorly suited for full-field imaging applications because their high spatial coherence leads to coherent artifacts known as speckle that corrupt image formation. We recently demonstrated that random lasers can be engineered to provide low spatial coherence. Here, we exploit the low spatial coherence of specifically-designed random lasers to perform speckle-free full-field imaging in the setting of significant optical scattering. We quantitatively demonstrate that images generated with random laser illumination exhibit higher resolution than images generated with spatially coherent illumination. By providing intense laser illumination without the drawback of coherent artifacts, random lasers are well suited for a host of full-field imaging applications from full-field microscopy to digital light projector systems.

X-ray absorption spectroscopy by full-field X-ray microscopy of a thin graphite flake: Imaging and electronic structure via the carbon K-edge

We demonstrate that near-edge X-ray-absorption fine-structure spectra combined with full-field transmission X-ray microscopy can be used to study the electronic structure of graphite flakes consisting of a few graphene layers. The flake was produced by exfoliation using sodium cholate and then isolated by means of density-gradient ultracentrifugation. An image sequence around the carbon K-edge, analyzed by using reference spectra for the in-plane and out-of-plane regions of the sample, is used to map and spectrally characterize the flat and folded regions of the flake. Additional spectral features in both π and σ regions are observed, which may be related to the presence of topological defects. Doping by metal impurities that were present in the original exfoliated graphite is indicated by the presence of a pre-edge signal at 284.2 eV.

Imaging cells and sub-cellular structures with ultrahigh resolution full-field X-ray microscopy

Our experimental results demonstrate that full-field hard-X-ray microscopy is finally able to investigate the internal structure of cells in tissues. This result was made possible by three main factors: the use of a coherent (synchrotron) source of X-rays, the exploitation of contrast mechanisms based on the real part of the refractive index and the magnification provided by high-resolution Fresnel zone-plate objectives. We specifically obtained high-quality microradiographs of human and mouse cells with 29nm Rayleigh spatial resolution and verified that tomographic reconstruction could be implemented with a final resolution level suitable for subcellular features. We also demonstrated that a phase retrieval method based on a wave propagation algorithm could yield good subcellular images starting from a series of defocused microradiographs. The concluding discussion compares cellular and subcellular hard-X-ray microradiology with other techniques and evaluates its potential impact on biomedical research.

Stagnation point flow of wormlike micellar solutions in a microfluidic cross-slot device: Effects of surfactant concentration and ionic environment

We employ the techniques of microparticle image velocimetry and full-field birefringence microscopy combined with mechanical measurements of the pressure drop to perform a detailed characterization of the extensional rheology and elastic flow instabilities observed for a range of wormlike micellar solutions flowing through a microfluidic cross-slot device. As the flow rate through the device is increased, the flow first bifurcates from a steady symmetric to a steady asymmetric configuration characterized by a birefringent strand of highly aligned micellar chains oriented along the shear-free centerline of the flow field. At higher flow rates the flow becomes three dimensional and time dependent and is characterized by aperiodic spatiotemporal fluctuations of the birefringent strand. The extensional properties and critical conditions for the onset of flow instabilities in the fluids are highly dependent on the fluid formulation (surfactant concentration and ionic strength) and the resulting changes in the linear viscoelasticity and nonlinear shear rheology of the fluids. By combining the measurements of critical conditions for the flow transitions with the viscometric material properties and the degree of shear-thinning characterizing each test fluid, it is possible to construct a stability diagram for viscoelastic flow of complex fluids in the cross-slot geometry.

Imaging at the Nanoscale With Practical Table-Top EUV Laser-Based Full-Field Microscopes

The demonstration of table-top high average power extreme-ultraviolet (EUV) lasers combined with the engineering of specialized optics has enabled the demonstration of full-field microscopes that have achieved tens of nanometer spatial resolution. This paper describes the geometry of the EUV microscopes tailored to specific imaging applications. The microscope illumination characteristics are assessed and an analysis on the microscope's spatial resolution is presented. Examples of the capabilities of these table-top EUV aerial microscopes for imaging nanostructures and surfaces are presented.

Symmetry breaking in the formation of magnetic vortex states in a permalloy nanodisk

The magnetic vortex in nanopatterned elements is currently attracting enormous interest. A priori, one would assume that the formation of magnetic vortex states should exhibit a perfect symmetry, because the magnetic vortex has four degenerate states. Here we show the first direct observation of an asymmetric phenomenon in the formation process of vortex states in a permalloy nanodisk using high-resolution full-field magnetic transmission soft X-ray microscopy. Micromagnetic simulations confirm that the intrinsic Dzyaloshinskii-Moriya interaction, which arises from the spin-orbit coupling due to the lack of inversion symmetry near the disk surface, as well as surface-related extrinsic factors, is decisive for the asymmetric formation of vortex states.

Extensional rheology and elastic instabilities of a wormlike micellar solution in a microfluidic cross-slot device

Wormlike micellar surfactant solutions are encountered in a wide variety of important applications, including enhanced oil recovery and ink-jet printing, in which the fluids are subjected to high extensional strain rates. In this contribution we present an experimental investigation of the flow of a model wormlike micellar solution (cetyl pyridinium chloride and sodium salicylate in deionised water) in a well-defined stagnation point extensional flow field generated within a microfluidic cross-slot device. We use micro-particle image velocimetry (μ-PIV) and full-field birefringence microscopy coupled with macroscopic measurements of the bulk pressure drop to make a quantitative characterization of the fluid's rheological response over a wide range of deformation rates. The flow field in the micromachined cross-slot is first characterized for viscous flow of a Newtonian fluid, and μ-PIV measurements show the flow field remains symmetric and stable up to moderately high Reynolds number, Re ≈ 20, and nominal strain rate, nom ≈ 635 s−1. By contrast, in the viscoelastic micellar solution the flow field remains symmetric only for low values of the strain rate such that nom ≤ λM−1, where λM = 2.5 s is the Maxwell relaxation time of the fluid. In this stable flow regime the fluid displays a localized and elongated birefringent strand extending along the outflow streamline from the stagnation point, and estimates of the apparent extensional viscosity can be obtained using the stress-optical rule and from the total pressure drop measured across the cross-slot channel. For moderate deformation rates (nom ≥ λM−1) the flow remains steady, but becomes increasingly asymmetric with increasing flow rate, eventually achieving a steady state of complete anti-symmetry characterized by a dividing streamline and birefringent strand connecting diagonally opposite corners of the cross-slot. Eventually, as the nominal imposed deformation rate is increased further, the asymmetric divided flow becomes time dependent. These purely elastic instabilities are reminiscent of those observed in cross-slot flows of polymer solutions, but seem to be strongly influenced by the effects of shear localization of the micellar fluid within the microchannels and around the re-entrant corners of the cross-slot.

X-ray mosaic nanotomography of large microorganisms

Full-field X-ray microscopy is a valuable tool for 3D observation of biological systems. In the soft X-ray domain organelles can be visualized in individual cells while hard X-ray microscopes excel in imaging of larger complex biological tissue. The field of view of these instruments is typically 10(3) times the spatial resolution. We exploit the assets of the hard X-ray sub-micrometer imaging and extend the standard approach by widening the effective field of view to match the size of the sample. We show that global tomography of biological systems exceeding several times the field of view is feasible also at the nanoscale with moderate radiation dose. We address the performance issues and limitations of the TOMCAT full-field microscope and more generally for Zernike phase contrast imaging. Two biologically relevant systems were investigated. The first being the largest known bacteria (Thiomargarita namibiensis), the second is a small myriapod species (Pauropoda sp.). Both examples illustrate the capacity of the unique, structured condenser based broad-band full-field microscope to access the 3D structural details of biological systems at the nanoscale while avoiding complicated sample preparation, or even keeping the sample environment close to the natural state.