Quantifying Embolism: Label-Free Volumetric Mapping of Thrombus Structure and Kinesis in a Microfluidic System with Optical Holography

Embolization of thrombotic material may lead to acute events such as ischemia and myocardial infarction. The embolus is the physical detachment from a primary thrombus that has developed under fluid shear rates. The physical characteristics (surface area coverage, volume, mass, and packing density) of a thrombus influence the overall flow dynamics of an occluding blood vessel. Here, the effectiveness of holographic quantitative phase microscopy (QPM) in identifying multiple morphological parameters of a thrombus (volume, surface area, and height) formed over collagen‐coated microfluidic channels by exerting a range of shear rates with anticoagulated platelet‐rich plasma (PRP) and whole blood is demonstrated. QPM enables the recording of entire thrombus volumes in real‐time using PRP and observed both growth and contraction trends of thrombi, without need for biochemical labeling. The process of emboli detachment in a microfluidic channel under pathophysiological shear rates (7500 and 12 500 s−1) is quantified. Rapid and direct quantification of an embolizing thrombus can enable the study of events during undesirable vessel occlusion and lead to targeting and early diagnosis of acute coronary and venous events.The authors received funding from the National Health and Medical Research Council of Australia and the Australian Research Council

Noise reduction in ultra-low light digital holographic microscopy using neural networks

Live cell imaging is challenging because the difficult balance of maintaining both cell viability and high signal to noise ratio throughout the entire imaging duration. Label free quantitative light microscopy techniques are powerful tools to image the volumetric activities in living cellular and sub-cellular biological systems, however there are minimal ways to identify phototoxicity. In this paper, we investigate the use of neural network to restore quantitative digital hologram micrographs at ultra-low light levels down to 0.06 /2 which approximately two orders of magnitude lower than sunlight. By developing an adaptive image restoration method specifically tailored for digital holograms, we demonstrated the 2x improvement in SSIM over existing denoising methods. This demonstration could open up new avenues for high resolution holographic microscopy using deep ultraviolet coherent sources and achieve high-resolution imaging with ultralow light illuminatio

Enhanced photoluminescence and reduced dimensionality via vacancy ordering in a 10H halide perovskite

H.L. would like to thank the University of St. Andrews for financial support via a St. Leonard’s scholarship. The authors acknowledge facilities access made possible through support from the EPSRC Light Element Analysis facility Grant EP/T019298/1 and the EPSRC Strategic Equipment Resource Grant EP/R023751/1.Vacancy-ordered halide perovskites have received great interest in optoelectronic applications. In this work, we report the novel inorganic halide Cs10MnSb6Cl30 with a distinctive 10H (10-layer hexagonal) perovskite polytype structure with (hcccc)2 stacking. Cs10MnSb6Cl30 has 30% B-site vacancies ordered at both corner- and face-sharing sites, resulting in [MnSb6Cl30]10–n columns, i.e., a reduction of octahedral connectivity to 1D. This results in enhanced photoluminescence in comparison to the previously reported 25% vacancy-ordered 3C polytype Cs4MnSb2Cl12 with 2D connectivity. This demonstrates not only the existence of the 10H perovskite structure in halides but also demonstrates the degree of B-site deficiency and stacking sequence variation as a direction to tune the optical properties of perovskite polytypes via vacancy rearrangements.Publisher PDFPeer reviewe

Quasiparticle thermal Hall angle and magnetoconductance in YBa_2Cu_3O_x

We present a way to extract the quasiparticle (qp) thermal conductivity Kappa_e and mean-free-path in YBa_2Cu_3O_x, using the thermal Hall effect and the magnetoconductance of Kappa_e. The results are very consistent with heat capacity experiments. Moreover, we find a simple relation between the thermal Hall angle Theta_Q and the H-dependence of Kappa_e, as well as numerical equality between Theta_Q and the electrical Hall angle. The findings also reveal an anomalously anisotropic scattering process in the normal state.Comment: 4 pages in Tex, 5 figures in EPS; replaced on 5/12/99, minor change

Improving focal photostimulation of cortical neurons with pre-derived wavefront correction

Recent progress in neuroscience to image and investigate brain function has been made possible by impressive developments in optogenetic and opto-molecular tools. Such research requires advances in optical techniques for the delivery of light through brain tissue with high spatial resolution. The tissue causes distortions to the wavefront of the incoming light which broadens the focus and consequently reduces the intensity and degrades the resolution. Such effects are detrimental in techniques requiring focal stimulation. Adaptive wavefront correction has been demonstrated to compensate for these distortions. However, iterative derivation of the corrective wavefront introduces time constraints that limit its applicability to probe living cells. Here, we demonstrate that we can pre-determine and generalize a small set of Zernike modes to correct for aberrations of the light propagating through specific brain regions. A priori identification of a corrective wavefront is a direct and fast technique that improves the quality of the focus without the need for iterative adaptive wavefront correction. We verify our technique by measuring the efficiency of two-photon photolysis of caged neurotransmitters along the dendrites of a whole-cell patched neuron. Our results show that encoding the selected Zernike modes on the excitation ight can improve light propagation through brain slices of rats as observed by the neuron’s evoked excitatory post-synaptic potential in response to localized focal uncaging at the spines of the neuron’s dendrites.This work is supported by the Australian Research Council Discovery Project (contract no. DP140101555) and National Health and Medical Research Council Project Grant (contract no. PG1105944)

Fibrin exposure triggers αIIbβ3-independent platelet aggregate formation, ADAM10 activity and glycoprotein VI shedding in a charge-dependent manner

Background Collagen and fibrin engagement and activation of glycoprotein (GP) VI induces proteolytic cleavage of the GPVI ectodomain generating shed soluble GPVI (sGPVI). Collagen‐mediated GPVI shedding requires intracellular signalling to release the sGPVI, mediated by A Disintegrin And Metalloproteinase 10 (ADAM10); however, the precise mechanism by which fibrin induces GPVI shedding remains elusive. Plasma sGPVI levels are elevated in patients with coagulopathies, sepsis, or inflammation and can predict onset of sepsis and sepsis‐related mortality; therefore, it is clinically important to understand the mechanisms of GPVI shedding under conditions of minimal collagen exposure. Objectives Our aim was to characterize mechanisms by which fibrin‐GPVI interactions trigger GPVI shedding. Methods Platelet aggregometry, sGPVI ELISA, and an ADAM10 fluorescence resonance energy transfer assay were used to measure fibrin‐mediated platelet responses. Results Fibrin induced αIIbβ3‐independent washed platelet aggregate formation, GPVI shedding, and increased ADAM10 activity, all of which were insensitive to pre‐treatment with inhibitors of Src family kinases but were divalent cation‐ and metalloproteinase‐dependent. In contrast, treatment of washed platelets with other GPVI ligands, collagen, and collagen‐related peptide caused αIIbβ3‐dependent platelet aggregation and GPVI release but did not increase constitutive ADAM10 activity. Conclusions Fibrin engages GPVI in a manner that differs from other GPVI ligands. Inclusion of polyanionic molecules disrupted fibrin‐induced platelet aggregate formation and sGPVI release, suggesting that electrostatic charge may play a role in fibrin/GPVI engagement. It may be feasible to exploit this property and specifically disrupt GPVI/fibrin interactions whilst sparing GPVI/collagen engagement.Fibrin engages GPVI in a manner that differs from other GPVI ligands. Inclusion of polyanionic molecules disrupted fibrin‐induced platelet aggregate formation and sGPVI release, suggesting that electrostatic charge may play a role in fibrin/GPVI engagement. It may be feasible to exploit this property and specifically disrupt GPVI/fibrin interactions whilst sparing GPVI/collagen engagement.National Health and Medical Research Council of Australia; Australian Research Council; THANZ Science and Education Research Gran

Compact flexible multi-pass rotary delay line using spinning micro-machined mirrors

We propose a new method to extend the path length tunability of rotary delay-lines. This method was shown to achieve a duty cycle of >80% and repetition rates of over 40 kHz. The new method relies on a new multi-segmented micro-machined mirror and serial injection of a single reflection onto separate segments of this mirror. The tunability is provided by the relative positioning of each reflective point on the mirror segments. There are two distinct modes of operation: synchronous and asynchronous. By simply manipulating the spatial position of the returning paths over the respective mirror segments, we can switch between increasing the repetition rate (asynchronous mode) or the total delay path (synchronous mode). We experimentally demonstrated up to 8 m/s scans with repetition rates of up to 42.7 kHz. Furthermore, we present numerical simulations of 18 reflection points to illustrate possibility of achieving a scan speed of up to 80 m/s. Through intermediate combinations of synchronous and asynchronous operation modes with 4 or more passes, we also show that the system can simultaneously increase both repetition rate and scan depth.Australian Research Council Early Career Researcher Award (DE160100843), Future Engineering Research Leadership Funds, Discovery Translation Funds

Where is the evidence for emergency planning: a scoping review

Background Recent terrorist attacks and natural disasters have led to an increased awareness of the importance of emergency planning. However, the extent to which emergency planners can access or use evidence remains unclear. The aim of this study was to identify, analyse and assess the location, source and quality of emergency planning publications in the academic and UK grey literature. Methods We conducted a scoping review, using as data sources for academic literature Embase, Medline, Medline in Process, Psychinfo, Biosis, Science Citation Index, Cinahl, Cochrane library and Clinicaltrials.gov. For grey literature identification we used databases at the Health Protection Agency, NHS Evidence, British Association of Immediate Care Schemes, Emergency Planning College and the Health and Safety Executive, and the websites of UK Department of Health Emergency Planning Division and UK Resilience. Aggregative synthesis was used to analyse papers and documents against a framework based on a modified FEMA Emergency Planning cycle. Results Of 2736 titles identified from the academic literature, 1603 were relevant. 45% were from North America, 27% were commentaries or editorials and 22% were event reports. Of 192 documents from the grey literature, 97 were relevant. 76% of these were event reports. The majority of documents addressed emergency planning and response. Very few documents related to hazard analysis, mitigation or capability assessment. Conclusions Although a large body of literature exists, its validity and generalisability is unclear There is little evidence that this potential evidence base has been exploited through synthesis to inform policy and practice. The type and structure of evidence that would be of most value of emergency planners and policymakers has yet to be identified

Modified inverted selective plane illumination microscopy for sub-micrometer imaging resolution in polydimethylsiloxane soft lithography devices

Moldable, transparent polydimethylsiloxane (PDMS) elastomer microdevices enable a broad range of complex studies of three-dimensional cellular networks in their microenvironment in vitro. However, the uneven distribution of refractive index change, external to PDMS devices and internally in the sample chamber, creates a significant optical path difference (OPD) that distorts the light sheet beam and so restricts diffraction limited performance. We experimentally showed that an OPD of 120 μm results in the broadening of the lateral point spread function by over 4-fold. In this paper, we demonstrate steps to adapt a commercial inverted selective plane illumination microscope (iSPIM) and remove the OPD so as to achieve sub-micrometer imaging ranging from 0.6 ± 0.04 μm to 0.91 ± 0.03 μm of a fluorescence biological sample suspended in regular saline (RI ≈1.34) enclosed in 1.2 to 2 mm thick micromolded PDMS microdevices. We have proven that the removal of the OPD from the external PDMS layer by refractive index (RI) matching with a readily accessible, inexpensive sucrose solution is critical to achieve a >3-fold imaging resolution improvement. To monitor the RI matching process, a single-mode fiber (SMF) illuminator was integrated into the iSPIM. To remove the OPD inside the PDMS channel, we used an electrically tunable lens (ETL) that par-focuses the light sheet beam with the detection objective lens and so minimised axial distortions to attain sub-micrometer imaging resolution. We termed this new light sheet imaging protocol as modified inverted selective plane illumination microscopy (m-iSPIM). Using the high spatial–temporal 3D imaging of m-iSPIM, we experimentally captured single platelet (≈2 μm) recruitment to a platelet aggregate (22.5 μm × 22.5 μm × 6 μm) under flow at a 150 μm depth within a microfluidic channel. m-iSPIM paves the way for the application of light sheet imaging to a wide range of 3D biological models in microfluidic devices which recapitulate features of the physiological microenvironment and elucidate subcellular responses.This work was supported by grants Australian Research Council (DP200100364, DP190100039, DE160100843) and ANU Major Equipment grant (15MEC36, 16MEC26) and additional funds from ARC Centre of Excellence Translational Photosynthesis

Different prion disease phenotypes result from inoculation of cattle with two temporally separated sources of sheep scrapie from Great Britain

BACKGROUND: Given the theoretical proposal that bovine spongiform encephalopathy (BSE) could have originated from sheep scrapie, this study investigated the pathogenicity for cattle, by intracerebral (i.c.) inoculation, of two pools of scrapie agents sourced in Great Britain before and during the BSE epidemic. Two groups of ten cattle were each inoculated with pools of brain material from sheep scrapie cases collected prior to 1975 and after 1990. Control groups comprised five cattle inoculated with sheep brain free from scrapie, five cattle inoculated with saline, and for comparison with BSE, naturally infected cattle and cattle i.c. inoculated with BSE brainstem homogenate from a parallel study. Phenotypic characterisation of the disease forms transmitted to cattle was conducted by morphological, immunohistochemical, biochemical and biological methods. RESULTS: Disease occurred in 16 cattle, nine inoculated with the pre-1975 inoculum and seven inoculated with the post-1990 inoculum, with four cattle still alive at 83 months post challenge (as at June 2006). The different inocula produced predominantly two different disease phenotypes as determined by histopathological, immunohistochemical and Western immunoblotting methods and biological characterisation on transmission to mice, neither of which was identical to BSE. Whilst the disease presentation was uniform in all scrapie-affected cattle of the pre-1975 group, the post-1990 inoculum produced a more variable disease, with two animals sharing immunohistochemical and molecular profile characteristics with animals in the pre-1975 group. CONCLUSION: The study has demonstrated that cattle inoculated with different pooled scrapie sources can develop different prion disease phenotypes, which were not consistent with the phenotype of BSE of cattle and whose isolates did not have the strain typing characteristics of the BSE agent on transmission to mice