Hollow Bessel-like beam as an optical guide for a stream of microscopic particles.

Current aerosol sample injection methods for coherent x-ray morphology suffer from excessive sample consumption due to the dispersion of the aerosol. To remedy this we propose here a high aspect ratio optical funnel by using a hollow Bessel-like beam with variable divergence, which may reduce sample consumption significantly. We present estimated optical forces exerted on the particles in the transverse plane, depending on various experimental conditions. We show that light pressure imposed by a funnel formed with 4.2 W continuous wave laser is sufficient to divert a stream of 2 µm polystyrene particles travelling ~50 m/s by ~1.5 × 10(-3) rad.

Imaging Platelet Processes and Function-Current and Emerging Approaches for Imaging in vitro and in vivo.

Platelets are small anucleate cells that are essential for many biological processes including hemostasis, thrombosis, inflammation, innate immunity, tumor metastasis, and wound healing. Platelets circulate in the blood and in order to perform all of their biological roles, platelets must be able to arrest their movement at an appropriate site and time. Our knowledge of how platelets achieve this has expanded as our ability to visualize and quantify discreet platelet events has improved. Platelets are exquisitely sensitive to changes in blood flow parameters and so the visualization of rapid intricate platelet processes under conditions found in flowing blood provides a substantial challenge to the platelet imaging field. The platelet's size (~2 µm), rapid activation (milliseconds), and unsuitability for genetic manipulation, means that appropriate imaging tools are limited. However, with the application of modern imaging systems to study platelet function, our understanding of molecular events mediating platelet adhesion from a single-cell perspective, to platelet recruitment and activation, leading to thrombus (clot) formation has expanded dramatically. This review will discuss current platelet imaging techniques in vitro and in vivo, describing how the advancements in imaging have helped answer/expand on platelet biology with a particular focus on hemostasis. We will focus on platelet aggregation and thrombus formation, and how platelet imaging has enhanced our understanding of key events, highlighting the knowledge gained through the application of imaging modalities to experimental models in vitro and in vivo. Furthermore, we will review the limitations of current imaging techniques, and questions in thrombosis research that remain to be addressed. Finally, we will speculate how the same imaging advancements might be applied to the imaging of other vascular cell biological functions and visualization of dynamic cell-cell interactions.

Fibrin exposure triggers αIIbß3-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 αIIbß3-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 αIIbß3-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.

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 light 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.

Ultrahigh-resolution optical coherence elastography through a micro-endoscope: towards in vivo imaging of cellular-scale mechanics.

In this paper, we describe a technique capable of visualizing mechanical properties at the cellular scale deep in living tissue, by incorporating a gradient-index (GRIN)-lens micro-endoscope into an ultrahigh-resolution optical coherence elastography system. The optical system, after the endoscope, has a lateral resolution of 1.6 µm and an axial resolution of 2.2 µm. Bessel beam illumination and Gaussian mode detection are used to provide an extended depth-of-field of 80 µm, which is a 4-fold improvement over a fully Gaussian beam case with the same lateral resolution. Using this system, we demonstrate quantitative elasticity imaging of a soft silicone phantom containing a stiff inclusion and a freshly excised malignant murine pancreatic tumor. We also demonstrate qualitative strain imaging below the tissue surface on in situ murine muscle. The approach we introduce here can provide high-quality extended-focus images through a micro-endoscope with potential to measure cellular-scale mechanics deep in tissue. We believe this tool is promising for studying biological processes and disease progression in vivo.