scholarly journals Lights, Camera, Path Splitter: A New Approach for Truly Simultaneous Dual Optical Mapping of the Heart with a Single Camera

2019 ◽  
Author(s):  
Rafael Jaimes ◽  
Damon McCullough ◽  
Bryan Siegel ◽  
Luther Swift ◽  
James Hiebert ◽  
...  
Keyword(s):  
Optical Mapping ◽  
Frame Rate ◽  
Small Rodent ◽  
Wide Field ◽  
Single Camera ◽  
Spatiotemporal Resolution ◽  
Cardiovascular Research ◽  
Excitation Light ◽  
Physiological Models ◽  
Dual Mapping

ABSTRACTBackgroundOptical mapping of transmembrane voltage and intracellular calcium is a powerful tool for investigating cardiac physiology and pathophysiology. However, simultaneous dual mapping of two fluorescent probes remains technically challenging. We introduce a novel, easy-to-use approach that requires a path splitter, single camera and excitation light to simultaneously acquire voltage and calcium signals from whole heart preparations, which can be applied to other physiological models – including neurons and isolated cardiomyocytes.ResultsComplementary probes were selected that could be excited with a single wavelength light source. Langendorff-perfused hearts (rat, swine) were stained and imaged using a sCMOS camera outfitted with an optical path splitter to simultaneously acquire two emission fields at high spatial and temporal resolution. Voltage (RH237) and calcium (Rhod2) signals were acquired concurrently on a single sensor, resulting in two 384×256 images at 814 frames per second. At this frame rate, the signal-to-noise ratio was 47 (RH237) and 85 (Rhod2). Imaging experiments were performed on small rodent hearts, as well as larger pig hearts with sufficient optical signals. In separate experiments, each dye was used independently to assess crosstalk and demonstrate signal specificity. Additionally, the effect of ryanodine on myocardial calcium transients was validated – with no measurable effect on the amplitude of optical action potentials. To demonstrate spatial resolution, ventricular tachycardia was induced –resulting in the novel finding that spatially discordant calcium alternans can be present in different regions of the heart, even when electrical alternans remain concordant. The described system excels in providing a wide field of view and high spatiotemporal resolution for a variety of cardiac preparations.ConclusionsWe report the first multiparametric mapping system that simultaneously acquires calcium and voltage signals from cardiac preparations, using a path splitter, single camera and excitation light. This approach eliminates the need for multiple cameras, excitation light patterning or frame interleaving. These features can aid in the adoption of dual mapping technology by the broader cardiovascular research community, and decrease the barrier of entry into panoramic heart imaging, as it reduces the number of required cameras.

scholarly journals Lights, camera, path splitter: a new approach for truly simultaneous dual optical mapping of the heart with a single camera

2019 ◽  
Vol 1 (1) ◽  
Author(s):  
Rafael Jaimes ◽  
Damon McCullough ◽  
Bryan Siegel ◽  
Luther Swift ◽  
James Hiebert ◽  
...  
Keyword(s):  
Optical Mapping ◽  
Frame Rate ◽  
Small Rodent ◽  
Wide Field ◽  
Single Camera ◽  
Spatiotemporal Resolution ◽  
Cardiovascular Research ◽  
Excitation Light ◽  
Physiological Models ◽  
Dual Mapping

Abstract Background Optical mapping of transmembrane voltage and intracellular calcium is a powerful tool for investigating cardiac physiology and pathophysiology. However, simultaneous dual mapping of two fluorescent probes remains technically challenging. We introduce a novel, easy-to-use approach that requires a path splitter, single camera and excitation light to simultaneously acquire voltage and calcium signals from whole heart preparations, which can be applied to other physiological models – including neurons and isolated cardiomyocytes. Results Complementary probes were selected that could be excited with a single wavelength light source. Langendorff-perfused hearts (rat, swine) were stained and imaged using a sCMOS camera outfitted with an optical path splitter to simultaneously acquire two emission fields at high spatial and temporal resolution. Voltage (RH237) and calcium (Rhod2) signals were acquired concurrently on a single sensor, resulting in two 384 × 256 images at 814 frames per second. At this frame rate, the signal-to-noise ratio was 47 (RH237) and 85 (Rhod2). Imaging experiments were performed on small rodent hearts, as well as larger pig hearts with sufficient optical signals. In separate experiments, each dye was used independently to assess crosstalk and demonstrate signal specificity. Additionally, the effect of ryanodine on myocardial calcium transients was validated – with no measurable effect on the amplitude of optical action potentials. To demonstrate spatial resolution, ventricular tachycardia was induced –resulting in the novel finding that spatially discordant calcium alternans can be present in different regions of the heart, even when electrical alternans remain concordant. The described system excels in providing a wide field of view and high spatiotemporal resolution for a variety of cardiac preparations. Conclusions We report the first multiparametric mapping system that simultaneously acquires calcium and voltage signals from cardiac preparations, using a path splitter, single camera and excitation light. This approach eliminates the need for multiple cameras, excitation light patterning or frame interleaving. These features can aid in the adoption of dual mapping technology by the broader cardiovascular research community, and decrease the barrier of entry into panoramic heart imaging, as it reduces the number of required cameras.

scholarly journals CHARACTERIZATION OF MODERN CCD AND CMOS SENSORS FOR SKY SURVEYS

2021 ◽  
Vol 53 ◽  
pp. 190-197
Author(s):  
S. Karpov ◽  
A. Christov ◽  
A. Bajat ◽  
R. Cunniffe ◽  
M. Prouza
Keyword(s):  
Laboratory Testing ◽  
Frame Rate ◽  
Wide Field ◽  
Large Format ◽  
Ccd Cameras ◽  
Sky Survey ◽  
Monitoring Applications ◽  
Cmos Sensors ◽  
Robotic Telescopes

Here we review the efforts we take in a newly established laboratory inside Institute of Physics in Prague in order to characterize modern large-format CCD and CMOS sensors for sky survey applications. While the laboratory is primarily established in order to participate in low-level CCD sensor characterization for LSST project, we also managed to perform a thorough laboratory testing of recently released Andor Marana sCMOS (which is especially interesting for wide-field sky monitoring applications due to its large format, backilluminated design, high achievable frame rate and low read-out noise), as well as detailed measurements of response non-linearity of Moravian Instruments G4-16000 CCD cameras (based on large-format Kodak KAF-16803 chip) used in several robotic telescopes. We briefly review the results acquired on these cameras, as well as hardware and software we developed for the laboratory.

scholarly journals Electro-optic imaging enables efficient wide-field fluorescence lifetime microscopy

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Adam J. Bowman ◽  
Brannon B. Klopfer ◽  
Thomas Juffmann ◽  
Mark A. Kasevich
Keyword(s):  
Temporal Resolution ◽  
Single Molecule ◽  
Fluorescence Lifetime ◽  
Collection Efficiency ◽  
Frame Rate ◽  
Optical Systems ◽  
Wide Field ◽  
Pockels Cell ◽  
Low Light ◽  
Optical Efficiency

Abstract Nanosecond temporal resolution enables new methods for wide-field imaging like time-of-flight, gated detection, and fluorescence lifetime. The optical efficiency of existing approaches, however, presents challenges for low-light applications common to fluorescence microscopy and single-molecule imaging. We demonstrate the use of Pockels cells for wide-field image gating with nanosecond temporal resolution and high photon collection efficiency. Two temporal frames are obtained by combining a Pockels cell with a pair of polarizing beam-splitters. We show multi-label fluorescence lifetime imaging microscopy (FLIM), single-molecule lifetime spectroscopy, and fast single-frame FLIM at the camera frame rate with 103–105 times higher throughput than single photon counting. Finally, we demonstrate a space-to-time image multiplexer using a re-imaging optical cavity with a tilted mirror to extend the Pockels cell technique to multiple temporal frames. These methods enable nanosecond imaging with standard optical systems and sensors, opening a new temporal dimension for wide-field low-light microscopy.

scholarly journals High-resolution single-camera photogrammetry: incorporation of refraction at a fluid interface

2019 ◽  
Vol 61 (1) ◽  
Author(s):  
A. S. González-Vera ◽  
T. J. S. Wilting ◽  
A. P. C. Holten ◽  
G. J. F. van Heijst ◽  
M. Duran-Matute
Keyword(s):  
Water Surface ◽  
Dimensional Space ◽  
Three Dimensional ◽  
Frame Rate ◽  
Dimensional Structure ◽  
Surface Plastic ◽  
Single Camera ◽  
Sediment Bed ◽  
Image Velocimetry ◽  
Resolution Single

Abstract Photogrammetry uses images of a three-dimensional structure to derive information about its shape and position. In this work, a photogrammetric technique is implemented with a single camera and a digital projector to measure changes in an underwater sediment bed. This implementation incorporates refraction at an interface allowing for measurements through a deformed or changing water surface. The digital projector provides flexibility in choosing projected patterns and has a high frame rate, which allows to easily increase the spatial and temporal resolution of the measurements. The technique requires first for both the camera and the projector to be calibrated using triangulation. With the calibration, we construct lines in three-dimensional space that originate from the projector and the camera, and intersect on the surface to be measured. To correctly incorporate refraction due to a change of medium, each line in space is recalculated from its intersection with the interface using Snell’s law. This has the benefit that only one calibration for measurements is needed if the location and shape of the interface are known. The technique is validated by measuring a submerged undulated surface, plastic objects and a sediment bed. In particular, the undulated plate is reconstructed under a flat and a parabolic water surface. Finally, the technique is used in combination with particle image velocimetry to dynamically measure a changing sediment bed under an oscillating flow and the flow velocity at the free surface. Graphic abstract

scholarly journals Extending resolution of structured illumination microscopy with sparse deconvolution

2021 ◽  
Author(s):  
Weisong Zhao ◽  
Shiqun Zhao ◽  
Liuju Li ◽  
Xiaoshuai Huang ◽  
Shijia Xing ◽  
...  
Keyword(s):  
Signal To Noise Ratio ◽  
Three Dimensional ◽  
Super Resolution ◽  
Live Cell ◽  
Frame Rate ◽  
Live Cells ◽  
Photon Flux ◽  
Structured Illumination ◽  
Structured Illumination Microscopy ◽  
Spatiotemporal Resolution

Abstract The spatial resolutions of live-cell super-resolution microscopes are limited by the maximum collected photon flux. Taking advantage of a priori knowledge of the sparsity and continuity of biological structures, we develop a deconvolution algorithm that further extends the resolution of super-resolution microscopes under the same photon budgets by nearly twofold. As a result, sparse structured illumination microscopy (Sparse-SIM) achieves ~60 nm resolution at a 564 Hz frame rate, allowing it to resolve intricate structural intermediates, including small vesicular fusion pores, ring-shaped nuclear pores formed by different nucleoporins, and relative movements between the inner and outer membranes of mitochondria in live cells. Likewise, sparse deconvolution can be used to increase the three-dimensional resolution and contrast of spinning-disc confocal-based SIM (SD-SIM), and operates under conditions with the insufficient signal-to-noise-ratio, all of which allows routine four-color, three-dimensional, ~90 nm resolution live-cell super-resolution imaging. Overall, sparse deconvolution may be a general tool to push the spatiotemporal resolution limits of live-cell fluorescence microscopy.

scholarly journals Stop the beat to see the rhythm: excitation-contraction uncoupling in cardiac research

2021 ◽  
Author(s):  
Luther M. Swift ◽  
Matthew W. Kay ◽  
Crystal M. Ripplinger ◽  
Nikki Gillum Posnack
Keyword(s):  
Electrical Activity ◽  
Cardiac Electrophysiology ◽  
Cardiac Tissue ◽  
Optical Mapping ◽  
Myocardial Contraction ◽  
Motion Artifacts ◽  
Secondary Effects ◽  
Metabolic Demand ◽  
Cardiovascular Research ◽  
Fluorescent Indicators

Optical mapping is an imaging technique that is extensively used in cardiovascular research, wherein parameter-sensitive fluorescent indicators are used to study the electrophysiology and excitation-contraction coupling of cardiac tissues. Despite the many benefits of optical mapping, eliminating motion artifacts within the optical signals is a major challenge, as myocardial contraction interferes with the faithful acquisition of action potentials and intracellular calcium transients. As such, excitation-contraction uncoupling agents are frequently used to reduce signal distortion by suppressing contraction. Compared to other uncoupling agents, blebbistatin is the most frequently used as it offers increased potency with minimal direct effects on cardiac electrophysiology. Nevertheless, blebbistatin may exert secondary effects on electrical activity, metabolism, and coronary flow, and the incorrect administration of blebbistatin to cardiac tissue can prove detrimental, resulting in erroneous interpretation of optical mapping results. In this "Getting It Right" perspective, we briefly review the literature regarding the use of blebbistatin in cardiac optical mapping experiments, highlight potential secondary effects of blebbistatin on cardiac electrical activity and metabolic demand, and conclude with the consensus of the authors on best practices for effectively using blebbistatin in optical mapping studies of cardiac tissue.

scholarly journals Ultrasound functional neuroimaging reveals propagation of task-related brain activity in behaving primates

2018 ◽  
Author(s):  
Alexandre Dizeux ◽  
Marc Gesnik ◽  
Harry Ahnine ◽  
Kevin Blaize ◽  
Fabrice Arcizet ◽  
...  
Keyword(s):  
Temporal Resolution ◽  
Large Scale ◽  
Functional Neuroimaging ◽  
Anterior Cingulate ◽  
Global Network ◽  
High Temporal Resolution ◽  
Wide Field ◽  
Spatiotemporal Resolution ◽  
Macro Scale ◽  
The Brain

ABSTRACTIn recent decades, neuroimaging has played an invaluable role in improving the fundamental understanding of the brain. At the macro scale, neuroimaging modalities such as MRI, EEG, and MEG, exploit a wide field of view to explore the brain as a global network of interacting regions. However, this comes at the price of either limited spatiotemporal resolution or limited sensitivity. At the micro scale, electrophysiology is used to explore the dynamic aspects of neuronal activity with a very high temporal resolution. However, this modality requires a statistical averaging of several tens of single task responses. A large-scale neuroimaging modality of sufficient spatial and temporal resolution and sensitivity to study brain region activation dynamically would open new territories of possibility in neuroscienceWe show that neurofunctional ultrasound imaging (fUS) is both able to assess brain activation during single cognitive tasks within superficial and deeper areas of the frontal cortex areas, and image the directional propagation of information within and between these regions. Equipped with an fUS device, two macaque rhesus monkeys were instructed before a stimulus appeared to rest (fixation) or to look towards (saccade) or away (antisaccade) from a stimulus. Our results identified an abrupt transient change in activity for all acquisitions in the supplementary eye field (SEF) when the animals were required to change a rule regarding the task cued by a stimulus. Simultaneous imaging in the anterior cingulate cortex and SEF revealed a time delay in the directional functional connectivity of 0.27 ± 0.07 s and 0.9 ± 0.2 s for animals S and Y, respectively. These results provide initial evidence that recording cerebral hemodynamics over large brain areas at a high spatiotemporal resolution and sensitivity with neurofunctional ultrasound can reveal instantaneous monitoring of endogenous brain signals and behavior.

scholarly journals Single-Camera Multi-Parametric Optical Mapping During Local Excitation of Isolated Rat Heart

2011 ◽  
Vol 100 (3) ◽  
pp. 318a
Author(s):  
Peter Lee ◽  
Christian Bollensdorff ◽  
Joseph P. Wuskell ◽  
Leslie M. Loew ◽  
Peter Kohl
Keyword(s):  
Rat Heart ◽  
Optical Mapping ◽  
Isolated Rat Heart ◽  
Single Camera ◽  
Local Excitation ◽  
Isolated Rat

Lensless On-Chip Fluorescent Imaging Over an Ultra Wide Field-of-View

2010 ◽  
Author(s):  
Ahmet F. Coskun ◽  
Ting-Wei Su ◽  
Aydogan Ozcan
Keyword(s):  
High Throughput ◽  
Imaging Modality ◽  
Scattered Light ◽  
Fluorescent Imaging ◽  
Field Of View ◽  
Fluorescent Detection ◽  
Fluorescent Light ◽  
Wide Field ◽  
Excitation Light ◽  
Wide Field Of View

We introduce a lensless high-throughput fluorescent detection modality that can simultaneously image micro-objects and labeled cells over an ultra-wide field-of-view (FOV) of ∼8cm2 without the use of any lenses, thin-film filters and mechanical scanners. This lensfree platform utilizes total-internal-reflection (TIR) to block the excitation light, and an inexpensive absorption filter to remove the weakly scattered light that does not obey TIR. The emitted fluorescent light from the objects is then detected on the same chip without the use of any lenses. A digital deconvolution algorithm is used to resolve overlapping fluorescent spots, enabling a resolution of ∼40–50 μm over the entire field-of-view. Such an ultra wide field-of-view lensfree fluorescent imaging modality might be very valuable for high-throughput screening applications as well as quantification of rare cells such as circulating tumor cells using ultra-large microfluidic devices.

scholarly journals High spatiotemporal resolution optoacoustic sensing with photothermally induced acoustic vibrations in optical fibres

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Yizhi Liang ◽  
Huojiao Sun ◽  
Linghao Cheng ◽  
Long Jin ◽  
Bai-Ou Guan
Keyword(s):  
High Performance ◽  
Frame Rate ◽  
Optical Fibres ◽  
Acoustic Vibrations ◽  
Label Free ◽  
Spatiotemporal Resolution ◽  
Spatially Resolved ◽  
Diffusion Dynamics ◽  
Wide Range ◽  
Surrounding Fluid

AbstractOptoacoustic vibrations in optical fibres have enabled spatially resolved sensing, but the weak electrostrictive force hinders their application. Here, we introduce photothermally induced acoustic vibrations (PTAVs) to realize high-performance fibre-based optoacoustic sensing. Strong acoustic vibrations with a wide range of axial wavenumbers kz are photothermally actuated by using a focused pulsed laser. The local transverse resonant frequency and loss coefficient can be optically measured by an intra-core acoustic sensor via spectral analysis. Spatially resolved sensing is further achieved by mechanically scanning the laser spot. The experimental results show that the PTAVs can be used to resolve the acoustic impedance of the surrounding fluid at a spatial resolution of approximately 10 μm and a frame rate of 50 Hz. As a result, PTAV-based optoacoustic sensing can provide label-free visualization of the diffusion dynamics in microfluidics at a higher spatiotemporal resolution.

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