16,065 research outputs found
Simultaneous whole-animal 3D-imaging of neuronal activity using light field microscopy
3D functional imaging of neuronal activity in entire organisms at single cell
level and physiologically relevant time scales faces major obstacles due to
trade-offs between the size of the imaged volumes, and spatial and temporal
resolution. Here, using light-field microscopy in combination with 3D
deconvolution, we demonstrate intrinsically simultaneous volumetric functional
imaging of neuronal population activity at single neuron resolution for an
entire organism, the nematode Caenorhabditis elegans. The simplicity of our
technique and possibility of the integration into epi-fluoresence microscopes
makes it an attractive tool for high-speed volumetric calcium imaging.Comment: 25 pages, 7 figures, incl. supplementary informatio
PhOTO Zebrafish: A Transgenic Resource for In Vivo Lineage Tracing during Development and Regeneration
Background: Elucidating the complex cell dynamics (divisions, movement, morphological changes, etc.) underlying embryonic development and adult tissue regeneration requires an efficient means to track cells with high fidelity in space and time. To satisfy this criterion, we developed a transgenic zebrafish line, called PhOTO, that allows photoconvertible optical tracking of nuclear and membrane dynamics in vivo.
Methodology: PhOTO zebrafish ubiquitously express targeted blue fluorescent protein (FP) Cerulean and photoconvertible FP Dendra2 fusions, allowing for instantaneous, precise targeting and tracking of any number of cells using Dendra2 photoconversion while simultaneously monitoring global cell behavior and morphology. Expression persists through adulthood, making the PhOTO zebrafish an excellent tool for studying tissue regeneration: after tail fin amputation and photoconversion of a ~100µm stripe along the cut area, marked differences seen in how cells contribute to the new tissue give detailed insight into the dynamic process of regeneration. Photoconverted cells that contributed to the regenerate were separated into three distinct populations corresponding to the extent of cell division 7 days after amputation, and a subset of cells that divided the least were organized into an evenly spaced, linear orientation along the length of the newly regenerating fin.
Conclusions/Significance: PhOTO zebrafish have wide applicability for lineage tracing at the systems-level in the early embryo as well as in the adult, making them ideal candidate tools for future research in development, traumatic injury and regeneration, cancer progression, and stem cell behavior
Advanced optical imaging in living embryos
Developmental biology investigations have evolved from static studies of embryo anatomy and into dynamic studies of the genetic and cellular mechanisms responsible for shaping the embryo anatomy. With the advancement of fluorescent protein fusions, the ability to visualize and comprehend how thousands to millions of cells interact with one another to form tissues and organs in three dimensions (xyz) over time (t) is just beginning to be realized and exploited. In this review, we explore recent advances utilizing confocal and multi-photon time-lapse microscopy to capture gene expression, cell behavior, and embryo development. From choosing the appropriate fluorophore, to labeling strategy, to experimental set-up, and data pipeline handling, this review covers the various aspects related to acquiring and analyzing multi-dimensional data sets. These innovative techniques in multi-dimensional imaging and analysis can be applied across a number of fields in time and space including protein dynamics to cell biology to morphogenesis
Do-It-Yourself Single Camera 3D Pointer Input Device
We present a new algorithm for single camera 3D reconstruction, or 3D input
for human-computer interfaces, based on precise tracking of an elongated
object, such as a pen, having a pattern of colored bands. To configure the
system, the user provides no more than one labelled image of a handmade
pointer, measurements of its colored bands, and the camera's pinhole projection
matrix. Other systems are of much higher cost and complexity, requiring
combinations of multiple cameras, stereocameras, and pointers with sensors and
lights. Instead of relying on information from multiple devices, we examine our
single view more closely, integrating geometric and appearance constraints to
robustly track the pointer in the presence of occlusion and distractor objects.
By probing objects of known geometry with the pointer, we demonstrate
acceptable accuracy of 3D localization.Comment: 8 pages, 6 figures, 2018 15th Conference on Computer and Robot Visio
Fast fluorescence microscopy for imaging the dynamics of embryonic development
Live imaging has gained a pivotal role in developmental biology since it increasingly allows real-time observation of cell behavior in intact organisms. Microscopes that can capture the dynamics of ever-faster biological events, fluorescent markers optimal for in vivo imaging, and, finally, adapted reconstruction and analysis programs to complete data flow all contribute to this success. Focusing on temporal resolution, we discuss how fast imaging can be achieved with minimal prejudice to spatial resolution, photon count, or to reliably and automatically analyze images. In particular, we show how integrated approaches to imaging that combine bright fluorescent probes, fast microscopes, and custom post-processing techniques can address the kinetics of biological systems at multiple scales. Finally, we discuss remaining challenges and opportunities for further advances in this field
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