Mark D. Junker

Whole-cell phase contrast imaging at the nanoscale using Fresnel Coherent Diffractive Imaging Tomography

X-ray tomography can provide structural information of whole cells in close to their native state. Radiation-induced damage, however, imposes a practical limit to image resolution, and as such, a choice between damage, image contrast, and image resolution must be made. New coherent diffractive imaging techniques, such Fresnel Coherent Diffractive Imaging (FCDI), allows quantitative phase information with exceptional dose efficiency, high contrast, and nano-scale resolution. Here we present three-dimensional quantitative images of a whole eukaryotic cell by FCDI at a spatial resolution below 70 nm with sufficient phase contrast to distinguish major cellular components. From our data, we estimate that the minimum dose required for a similar resolution is close to that predicted by the Rose criterion, considerably below accepted estimates of the maximum dose a frozen-hydrated cell can tolerate. Based on the dose efficiency, contrast, and resolution achieved, we expect this technique will find immediate applications in tomographic cellular characterisation.

Rapid, low dose X-ray diffractive imaging of the malaria parasite Plasmodium falciparum.

Phase-diverse X-ray coherent diffractive imaging (CDI) provides a route to high sensitivity and spatial resolution with moderate radiation dose. It also provides a robust solution to the well-known phase-problem, making on-line image reconstruction feasible. Here we apply phase-diverse CDI to a cellular sample, obtaining images of an erythrocyte infected by the sexual stage of the malaria parasite, Plasmodium falciparum, with a radiation dose significantly lower than the lowest dose previously reported for cellular imaging using CDI. The high sensitivity and resolution allow key biological features to be identified within intact cells, providing complementary information to optical and electron microscopy. This high throughput method could be used for fast tomographic imaging, or to generate multiple replicates in two-dimensions of hydrated biological systems without freezing or fixing. This work demonstrates that phase-diverse CDI is a valuable complementary imaging method for the biological sciences and ready for immediate application.

A soft X-ray beamline for quantitative nanotomography using ptychography

Soft X-ray nanotomography using ptychography allows quantitative imaging of the internal structure of biological and materials samples with high sensitivity. In this work, we describe progress toward the implementation of an interferometer-controlled microscope located at a beamline that provides coherent ux over the photon energy range of 200 to 2000 eV. Recent experimental results are presented to illustrate the potential for two- and three-dimensional imaging at the nanoscale.

Molar concentration from sequential 2-D water-window X-ray ptychography and X-ray fluorescence in hydrated cells

Recent developments in biological X-ray microscopy have allowed structural information and elemental distribution to be simultaneously obtained by combining X-ray ptychography and X-ray fluorescence microscopy. Experimentally, these methods can be performed simultaneously; however, the optimal conditions for each measurement may not be compatible. Here, we combine two distinct measurements of ultrastructure and elemental distribution, with each measurement performed under optimised conditions. By combining optimised ptychography and fluorescence information we are able to determine molar concentrations from two-dimensional images, allowing an investigation into the interactions between the environment sensing filopodia in fibroblasts and extracellular calcium. Furthermore, the biological ptychography results we present illustrate a point of maturity where the technique can be applied to solve significant problems in structural biology.

Shedding light on electrodeposition dynamics tracked in situ via soft X-ray coherent diffraction imaging

The in situ physicochemical analysis of nanostructured functional materials is crucial for advances in their design and production. X-ray coherent diffraction imaging (CDI) methods have recently demonstrated impressive potential for characterizing such materials with a high spatial resolution and elemental sensitivity; however, moving from the current ex situ static regime to the in situ dynamic one remains a challenge. By combining soft X-ray ptychography and single-shot keyhole CDI, we performed the first in situ spatiotemporal study on an electrodeposition process in a sealed wet environment, employed for the fabrication of oxygen-reduction catalysts, which are key components for alkaline fuel cells and metal-air batteries. The results provide the first experimental demonstration of theoretically predicted Turing–Hopf electrochemical pattern formation resulting from morphochemical coupling, adding a new dimension for the in-depth in situ characterization of electrodeposition processes in space and time.

Fresnel coherent diffractive imaging tomography of whole cells in capillaries

X-ray tomography can be used to study the structure of whole cells in close to their native state. Ptychographic Fresnel coherent diffractive imaging (FCDI) holds particular promise for high-resolution tomographic imaging with quantitative phase sensitivity. To avoid the common missing wedge problem in tomography, cells can be mounted in thin glass capillaries that allow access to the full 180° angular field. However, soft x-rays, which are preferred for cellular imaging, interact strongly with capillaries, sometimes leading to violation of the usual assumptions for coherent diffractive imaging (CDI) and introducing artifacts (i.e., phase wrapping) in the reconstructed images. Here, we describe a

A universal measure for coherence requirements in diffractive imaging

The requirements on the spatial and temporal coherence for conventional Coherent Diffractive Imaging (CDI) have been well-established in the literature based on Shannon sampling of the diffracted intensities. The spatial coherence length of the illumination must be larger than twice the lateral dimensions of the sample whilst the temporal coherence length must be larger than the maximum optical path length difference between the two edges of the sample for the highest order diffraction peaks. However, recent approaches to CDI which have included knowledge of the spatial and temporal coherence information in the image reconstruction have allowed us to relax these conventional coherence constraints, extending the applicability of the technique to less coherent sources. In light of these developments it is useful to revisit the idea of a coherence limit in partially coherent CDI and establish a ‘universal’ limit on the partial coherence that can be tolerated without any loss of information. In this paper we present a simple and straightforward description of the limit of spatial and temporal coherence in partially coherent CDI.

Partial Coherence: a Route to Performing Faster Coherent Diffraction Imaging

Coherent diffraction imaging (CDI) typically requires that the light source should be highly coherent both laterally and longitudinally. Beamlines at synchrotrons usually install a monochromator and slits to achieve a highly coherent source, leading to a large reduction of beam flux. We demonstrate that lateral and longitudinal partial coherence can be successfully included in a CDI reconstruction algorithm simultaneously, reducing the associated exposure time by two orders of magnitude. For the experimental case we present this allows the acquisition of CDI data in just 5 seconds compared to 20 minutes for full coherence. This significantly reduces the requirements on the stability of the imaging system as well as providing a route to imaging samples in real-time.