8,064 research outputs found
Time-Lapse Microscopy
Time-lapse microscopy is a powerful, versatile and constantly developing tool for real-time imaging of living cells. This review outlines the advances of time-lapse microscopy and refers to the most interesting reports, thus pointing at the fact that the modern biology and medicine are entering the thrilling and promising age of molecular cinematography
The evolution of the axonal transport toolkit
Neurons are highly polarized cells that critically depend on longârange, bidirectional transport between the cell body and synapse for their function. This continual and highly coordinated trafficking process, which takes place via the axon, has fascinated researchers since the early 20th century. Ramon y Cajal first proposed the existence of axonal trafficking of biological material after observing that dissociation of the axon from the cell body led to neuronal degeneration. Since these first indirect observations, the field has come a long way in its understanding of this fundamental process. However, these advances in our knowledge have been aided by breakthroughs in other scientific disciplines, as well as the parallel development of novel tools, techniques and model systems. In this review, we summarize the evolution of tools used to study axonal transport and discuss how their deployment has refined our understanding of this process. We also highlight innovative tools currently being developed and how their addition to the available axonal transport toolkit might help to address key outstanding questions
The multiple faces of leukocyte interstitial migration
Spatiotemporal control of leukocyte dynamics within tissues is critical for successful innate and adaptive immune responses. Homeostatic trafficking and coordinated infiltration into and within sites of inflammation and infection rely on signaling in response to extracellular cues that in turn controls a variety of intracellular protein networks regulating leukocyte motility, migration, chemotaxis, positioning, and cellâcell interaction. In contrast to mesenchymal cells, leukocytes migrate in an amoeboid fashion by rapid cycles of actin polymerization and actomyosin contraction, and their migration in tissues is generally referred to as low adhesive and nonproteolytic. The interplay of actin network expansion, contraction, and adhesion shapes the exact mode of amoeboid migration, and in this review, we explore how leukocyte subsets potentially harness the same basic biomechanical mechanisms in a cell-type-specific manner. Most of our detailed understanding of these processes derives from in vitro migration studies in three-dimensional gels and confined spaces that mimic geometrical aspects of physiological tissues. We summarize these in vitro results and then critically compare them to data from intravital imaging of leukocyte interstitial migration in mouse tissues. We outline the technical challenges of obtaining conclusive mechanistic results from intravital studies, discuss leukocyte migration strategies in vivo, and present examples of mode switching during physiological interstitial migration. These findings are also placed in the context of leukocyte migration defects in primary immunodeficiencies. This overview of both in vitro and in vivo studies highlights recent progress in understanding the molecular and biophysical mechanisms that shape robust leukocyte migration responses in physiologically complex and heterogeneous environments
Quantum metrology and its application in biology
Quantum metrology provides a route to overcome practical limits in sensing
devices. It holds particular relevance to biology, where sensitivity and
resolution constraints restrict applications both in fundamental biophysics and
in medicine. Here, we review quantum metrology from this biological context,
focusing on optical techniques due to their particular relevance for biological
imaging, sensing, and stimulation. Our understanding of quantum mechanics has
already enabled important applications in biology, including positron emission
tomography (PET) with entangled photons, magnetic resonance imaging (MRI) using
nuclear magnetic resonance, and bio-magnetic imaging with superconducting
quantum interference devices (SQUIDs). In quantum metrology an even greater
range of applications arise from the ability to not just understand, but to
engineer, coherence and correlations at the quantum level. In the past few
years, quite dramatic progress has been seen in applying these ideas into
biological systems. Capabilities that have been demonstrated include enhanced
sensitivity and resolution, immunity to imaging artifacts and technical noise,
and characterization of the biological response to light at the single-photon
level. New quantum measurement techniques offer even greater promise, raising
the prospect for improved multi-photon microscopy and magnetic imaging, among
many other possible applications. Realization of this potential will require
cross-disciplinary input from researchers in both biology and quantum physics.
In this review we seek to communicate the developments of quantum metrology in
a way that is accessible to biologists and biophysicists, while providing
sufficient detail to allow the interested reader to obtain a solid
understanding of the field. We further seek to introduce quantum physicists to
some of the central challenges of optical measurements in biological science.Comment: Submitted review article, comments and suggestions welcom
Background-deflection Brillouin microscopy reveals altered biomechanics of intracellular stress granules by ALS protein FUS
Altered cellular biomechanics have been implicated as key photogenic triggers in age-related diseases. An aberrant liquid-to-solid phase transition, observed in in vitro reconstituted droplets of FUS protein, has been recently proposed as a possible pathogenic mechanism for amyotrophic lateral sclerosis (ALS). Whether such transition occurs in cell environments is currently unknown as a consequence of the limited measuring capability of the existing techniques, which are invasive or lack of subcellular resolution. Here we developed a non-contact and label-free imaging method, named background-deflection Brillouin microscopy, to investigate the three-dimensional intracellular biomechanics at a sub-micron resolution. Our method exploits diffraction to achieve an unprecedented 10,000-fold enhancement in the spectral contrast of single-stage spectrometers, enabling, to the best of our knowledge, the first direct biomechanical analysis on intracellular stress granules containing ALS mutant FUS protein in fixed cells. Our findings provide fundamental insights on the critical aggregation step underlying the neurodegenerative ALS disease
Development of an integrated opto-electric biosensor to dynamically examine cytometric proliferation and cytotoxicity
My doctoral research has focused on the development of microscale optical techniques for examining micro/bio fluidics. Preliminary work measured the velocity field in a microchannel, by optical slicing, using Confocal Laser Scanning Microscopy (CLSM). Next, Optical Serial Sectioning Microscopy (OSSM) was applied to examine thermometry by detecting the free Brownian motion of nano-particles suspended in mediums at different temperatures. An extension of this work used objective-based Total Internal Reflection Fluorescence Microscopy (TIRFM) to examine the hindered Brownian motion of nano-particles that were very close to a solid surface (within 1 mm).
An optically transparent and electrically conductive Indium Tin Oxide (ITO) biosensor and an integrated dynamic live cell imaging system were developed to dynamically examine changes in cell coverage area, cell morphology, cell-substrate adhesion, and cell-cell interaction. To our knowledge this is the first sensor capable of conducting simultaneous optical and electrical measurements. This system consists of an incubator, which keeps cells viable by providing the necessary environmental conditions (37 °C temperature and 5 % CO2), and multiple microscopy techniques, including multispectrum Interference Reflection Microscopy (MS-IRM), TIRFM, Epi-fluorescence Microscopy, Phase Contrast Microscopy (PCM), and Differential Interference Contrast Microscopy (DICM). Along with investigations of cytometric proliferation including cellular barrier functions, in vitro cytotoxicity experiments were also conducted to examine the effect of a drug (cytochalasin D, a toxic agent) on cellular motility and cellular morphology. These cytotoxicity results give us a fundamental understanding of the cellular processes induced by the drug, which will be invaluable in the search for methods of preventing metastases. In this research, MS-IRM is used to examine the focal contacts and the gap morphology between cells and substrates, DICM is used to examine the coverage area of cells, and impedance measurements are used to correlate these two parameters.
Advances in the understanding of vascular bio-transport in endothelial cells will have an impact on many aspects of cell biology, tissue engineering, and pharmacology. Particularly important will be the ability to test the popular hypothesis that the cell barrier function is regulated by specific cytoskeleton elements controlling intercellular and extracellular coupling
Polarized light microscopy in reproductive and developmental biology
Author Posting. © The Author(s), 2013. This is the author's version of the work. It is posted here by permission of John Wiley & Sons for personal use, not for redistribution. The definitive version was published in Molecular Reproduction and Development (2013), doi:10.1002/mrd.22221.The polarized light microscope reveals orientational order in native molecular structures inside
living cells, tissues, and whole organisms. Therefore, it is a powerful tool to monitor and analyze
the early developmental stages of organisms that lend themselves to microscopic observations. In
this article we briefly discuss the components specific to a traditional polarizing microscope and
some historically important observations on chromosome packing in sperm head, first zygote
division of the sea urchin, and differentiation initiated by the first uneven cell division in the
sand dollar. We then introduce the LC-PolScope and describe its use for measuring birefringence
and polarized fluorescence in living cells and tissues. Applications range from the enucleation of
mouse oocytes to analyzing the polarized fluorescence of the water strider acrosome. We end by
reporting first results on the birefringence of the developing chick brain, which we analyzed
between developmental stages of days 12 through 20.This work was supported by funds from the National Institute of General Medical Sciences
(grant 1R01GM100160-01A1 awarded to TT) and the National Institute of Biomedical Imaging
and Bioengineering (grant EB002045 awarded to RO)
Living cells and dynamic molecules observed with the polarized light microscope : the legacy of Shinya Inoué
Author Posting. © Marine Biological Laboratory, 2016. This article is posted here by permission of Marine Biological Laboratory for personal use, not for redistribution. The definitive version was published in Biological Bulletin 231 (2016): 85-95.In 1948, Shinya Inoué arrived in the United States for graduate studies at Princeton. A year later he came to Woods Hole, starting a long tradition of summer research at the Marine Biological Laboratory (MBL), which quickly became Inoué's scientific home. Primed by his Japanese mentor, Katsuma Dan, Inoué followed Dan's mantra to work with healthy, living cells, on a fundamental problem (mitosis), with a unique tool set that he refined for precise and quantitative observations (polarized light microscopy), and a fresh and brilliant mind that was unafraid of challenging current dogma. Building on this potent combination, Inoué contributed landmark observations and concepts in cell biology, including the notion that there are dynamic, fine structures inside living cells, in which molecular assemblies such as mitotic spindle fibers exist in delicate equilibrium with their molecular building blocks suspended in the cytoplasm. In the late 1970s and 1980s, Inoué and others at the MBL were instrumental in conceiving video microscopy, a groundbreaking technique which married light microscopy and electronic imaging, ushering in a revolution in how we know and what we know about living cells and the molecular mechanisms of life. Here, we recount some of Inoué's accomplishments and describe how his legacy has shaped current activities in polarized light imaging at the MBL.Preparation of this manuscript
was supported by grants from the National Institutes
of Health (no. GM100160 to TT; no. GM101701 to MS; and
no. GM114274 to RO); and by the Marine Biological Laboratory
start-up funds from the InoueÂŽ Family Endowment,
to TT
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