scholarly journals Quantitative linear dichroism imaging of molecular processes in living cells made simple by open software tools

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Alexey Bondar ◽  
Olga Rybakova ◽  
Josef Melcr ◽  
Jan Dohnálek ◽  
Petro Khoroshyy ◽  
...  
Keyword(s):  
Biological Systems ◽  
Linear Dichroism ◽  
Quantitative Information ◽  
Living Cells ◽  
Software Tools ◽  
Model Systems ◽  
Membrane Properties ◽  
Polarization Microscopy ◽  
Molecular Processes ◽  
Wide Range

AbstractFluorescence-detected linear dichroism microscopy allows observing various molecular processes in living cells, as well as obtaining quantitative information on orientation of fluorescent molecules associated with cellular features. Such information can provide insights into protein structure, aid in development of genetically encoded probes, and allow determinations of lipid membrane properties. However, quantitating and interpreting linear dichroism in biological systems has been laborious and unreliable. Here we present a set of open source ImageJ-based software tools that allow fast and easy linear dichroism visualization and quantitation, as well as extraction of quantitative information on molecular orientations, even in living systems. The tools were tested on model synthetic lipid vesicles and applied to a variety of biological systems, including observations of conformational changes during G-protein signaling in living cells, using fluorescent proteins. Our results show that our tools and model systems are applicable to a wide range of molecules and polarization-resolved microscopy techniques, and represent a significant step towards making polarization microscopy a mainstream tool of biological imaging.

scholarly journals Quantitative Linear Dichroism Imaging of Molecular Processes in Living Cells Made Simple by Open Software Tools

2020 ◽  
Author(s):  
Alexey Bondar ◽  
Olga Rybakova ◽  
Josef Melcr ◽  
Jan Dohnálek ◽  
Petro Khoroshyy ◽  
...  
Keyword(s):  
Biological Systems ◽  
Linear Dichroism ◽  
Quantitative Information ◽  
Living Cells ◽  
Software Tools ◽  
Model Systems ◽  
Membrane Properties ◽  
Polarization Microscopy ◽  
Molecular Processes ◽  
Wide Range

Abstract Fluorescence-detected linear dichroism microscopy allows observing various molecular processes in living cells, as well as obtaining quantitative information on orientation of fluorescent molecules associated with cellular features. Such information can provide insights into protein structure, aid in development of genetically encoded probes, and allow determinations of lipid membrane properties. However, quantitating and interpreting linear dichroism in biological systems has been laborious and unreliable. Here we present a set of open source ImageJ-based software tools that allow fast and easy linear dichroism visualization and quantitation, as well as extraction of quantitative information on molecular orientations, even in living systems. The tools were tested on model synthetic lipid vesicles, and applied to a variety of biological systems, including observations of conformational changes during G-protein signaling in living cells, using fluorescent proteins. Our results show that our tools and model systems are applicable to a wide range of molecular systems and polarization-resolved microscopy techniques, and represent a significant step towards making polarization microscopy a mainstream tool of biological imaging.

scholarly journals Giant unilamellar vesicles - a perfect tool to visualize phase separation and lipid rafts in model systems.

2009 ◽  
Vol 56 (1) ◽  
Author(s):  
Olga Wesołowska ◽  
Krystyna Michalak ◽  
Jadwiga Maniewska ◽  
Andrzej B Hendrich
Keyword(s):  
Phase Separation ◽  
Lipid Membranes ◽  
Average Diameter ◽  
Model Systems ◽  
Membrane Properties ◽  
Lipid Phase ◽  
Giant Unilamellar Vesicles ◽  
Unilamellar Vesicles ◽  
Simple Method ◽  
Wide Range

Model systems such as black lipid membranes or conventional uni- or multilamellar liposomes are commonly used to study membrane properties and structure. However, the construction and dimensions of these models excluded their direct optical microscopic observation. Since the introduction of the simple method of liposome electroformation in alternating electric field giant unilamellar vesicles (GUVs) have become an important model imitating biological membranes. Due to the average diameter of GUVs reaching up to 100 microm, they can be easily observed under a fluorescent or confocal microscope provided that the appropriate fluorescent probe was incorporated into the lipid phase during vesicle formation. GUVs can be formed from different lipid mixtures and they are stable in a wide range of physical conditions such as pH, pressure or temperature. This mini-review presents information about the methods of GUV production and their usage. Particularly, the use of GUVs in studying lipid phase separation and the appearance and behavior of lipid domains (rafts) in membranes is discussed but also other examples of GUVs use in membrane research are given. The experience of the authors in setting up the GUV-forming equipment and production of GUVs is also presented.

scholarly journals KymographClear and KymographDirect: two tools for the automated quantitative analysis of molecular and cellular dynamics using kymographs

2016 ◽  
Vol 27 (12) ◽  
pp. 1948-1957 ◽  
Author(s):  
Pierre Mangeol ◽  
Bram Prevo ◽  
Erwin J. G. Peterman
Keyword(s):  
Live Cell Imaging ◽  
Full Range ◽  
Simulated Data ◽  
Quantitative Information ◽  
Software Tools ◽  
Dynamic Processes ◽  
Color Coding ◽  
Wide Range ◽  
Automated Tools ◽  
User Friendly

Dynamic processes are ubiquitous and essential in living cells. To properly understand these processes, it is imperative to measure them in a time-dependent way and analyze the resulting data quantitatively, preferably with automated tools. Kymographs are single images that represent the motion of dynamic processes and are widely used in live-cell imaging. Although they contain the full range of dynamics, it is not straightforward to extract this quantitative information in a reliable way. Here we present two complementary, publicly available software tools, KymographClear and KymographDirect, that have the power to reveal detailed insight in dynamic processes. KymographClear is a macro toolset for ImageJ to generate kymographs that provides automatic color coding of the different directions of movement. KymographDirect is a stand-alone tool to extract quantitative information from kymographs obtained from a wide range of dynamic processes in an automated way, with high accuracy and reliability. We discuss the concepts behind these software tools, validate them using simulated data, and test them on experimental data. We show that these tools can be used to extract motility parameters from a diverse set of cell-biological experiments in an automated and user-friendly way.

Modulated polarization microscopy displays the dynamics of cytoskeletal structures in living, motile cells

1995 ◽  
Vol 53 ◽  
pp. 902-903
Author(s):  
J. R. Kuhn ◽  
M. Poenie
Keyword(s):  
Mitotic Spindle ◽  
Actin Filaments ◽  
Cell Shape ◽  
Dynamic Structure ◽  
Living Cells ◽  
Cytoskeletal Proteins ◽  
Polarization Microscopy ◽  
Video Enhancement ◽  
Ultrastructural Studies ◽  
Motile Cells

Cell shape and movement are controlled by elements of the cytoskeleton including actin filaments an microtubules. Unfortunately, it is difficult to visualize the cytoskeleton in living cells and hence follow it dynamics. Immunofluorescence and ultrastructural studies of fixed cells while providing clear images of the cytoskeleton, give only a static picture of this dynamic structure. Microinjection of fluorescently Is beled cytoskeletal proteins has proved useful as a way to follow some cytoskeletal events, but long terry studies are generally limited by the bleaching of fluorophores and presence of unassembled monomers.Polarization microscopy has the potential for visualizing the cytoskeleton. Although at present, it ha mainly been used for visualizing the mitotic spindle. Polarization microscopy is attractive in that it pro vides a way to selectively image structures such as cytoskeletal filaments that are birefringent. By combing ing standard polarization microscopy with video enhancement techniques it has been possible to image single filaments. In this case, however, filament intensity depends on the orientation of the polarizer and analyzer with respect to the specimen.

Microtubule Dynamics and Organelle Transport in Reticulomyxa

1990 ◽  
Vol 48 (3) ◽  
pp. 194-195
Author(s):  
Yih-Tai Chen ◽  
Ursula Euteneuer ◽  
Ken B. Johnson ◽  
Michael P. Koonce ◽  
Manfred Schliwa
Keyword(s):  
Plasma Membrane ◽  
Light Microscopy ◽  
Cytoplasmic Dynein ◽  
Living Cells ◽  
Microtubule Dynamics ◽  
Model Systems ◽  
Organelle Transport ◽  
Biochemical Characteristics ◽  
Dependent Transport

The application of video techniques to light microscopy and the development of motility assays in reactivated or reconstituted model systems rapidly advanced our understanding of the mechanism of organelle transport and microtubule dynamics in living cells. Two microtubule-based motors have been identified that are good candidates for motors that drive organelle transport: kinesin, a plus end-directed motor, and cytoplasmic dynein, which is minus end-directed. However, the evidence that they do in fact function as organelle motors is still indirect.We are studying microtubule-dependent transport and dynamics in the giant amoeba, Reticulomyxa. This cell extends filamentous strands backed by an extensive array of microtubules along which organelles move bidirectionally at up to 20 μm/sec (Fig. 1). Following removal of the plasma membrane with a mild detergent, organelle transport can be reactivated by the addition of ATP (1). The physiological, pharmacological and biochemical characteristics show the motor to be a cytoplasmic form of dynein (2).

scholarly journals Engineering the Interface between Inorganic Nanoparticles and Biological Systems through Ligand Design

Nanomaterials ◽  
2021 ◽  
Vol 11 (4) ◽  
pp. 1001
Author(s):  
Rui Huang ◽  
David C. Luther ◽  
Xianzhi Zhang ◽  
Aarohi Gupta ◽  
Samantha A. Tufts ◽  
...  
Keyword(s):  
Organic Synthesis ◽  
Molecular Design ◽  
Biological Systems ◽  
Ligand Design ◽  
Inorganic Nanoparticles ◽  
Organic Ligands ◽  
Biological Applications ◽  
Wide Range ◽  
Insight Into

Nanoparticles (NPs) provide multipurpose platforms for a wide range of biological applications. These applications are enabled through molecular design of surface coverages, modulating NP interactions with biosystems. In this review, we highlight approaches to functionalize nanoparticles with ”small” organic ligands (Mw < 1000), providing insight into how organic synthesis can be used to engineer NPs for nanobiology and nanomedicine.

scholarly journals Regulation of Cytochrome c Oxidase by Natural Compounds Resveratrol, (–)-Epicatechin, and Betaine

Cells ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 1346
Author(s):  
Icksoo Lee
Keyword(s):  
Cytochrome C ◽  
Cytochrome C Oxidase ◽  
Model Systems ◽  
Health Improvement ◽  
Published Data ◽  
Mitochondrial Electron Transport Chain ◽  
Naturally Occurring ◽  
Transport Chain ◽  
Wide Range ◽  
Experimental Model Systems

Numerous naturally occurring molecules have been studied for their beneficial health effects. Many compounds have received considerable attention for their potential medical uses. Among them, several substances have been found to improve mitochondrial function. This review focuses on resveratrol, (–)-epicatechin, and betaine and summarizes the published data pertaining to their effects on cytochrome c oxidase (COX) which is the terminal enzyme of the mitochondrial electron transport chain and is considered to play an important role in the regulation of mitochondrial respiration. In a variety of experimental model systems, these compounds have been shown to improve mitochondrial biogenesis in addition to increased COX amount and/or its enzymatic activity. Given that they are inexpensive, safe in a wide range of concentrations, and effectively improve mitochondrial and COX function, these compounds could be attractive enough for possible therapeutic or health improvement strategies.

scholarly journals Continuum modeling perspectives of non-Fourier heat conduction in biological systems

2021 ◽  
Vol 0 (0) ◽  
Author(s):  
Ákos Sudár ◽  
Gergely Futaki ◽  
Róbert Kovács
Keyword(s):  
Biological Systems ◽  
Phase Lag ◽  
Dual Phase ◽  
Continuum Modeling ◽  
Biological Origin ◽  
Ultrasound Surgery ◽  
Thermal Models ◽  
Wide Range ◽  
Fourier Heat Conduction ◽  
The Continuum

Abstract The thermal modeling of biological systems is increasingly important in the development of more advanced and more precise techniques such as ultrasound surgery. One of the primary barriers is the complexity of biological materials: the geometrical, structural, and material properties vary in a wide range. In the present paper, we focus on the continuum modeling of heterogeneous materials of biological origin. There are numerous examples in the literature for non-Fourier thermal models. However, as we realized, they are associated with a few common misconceptions. Therefore, we first aim to clarify the basic concepts of non-Fourier thermal models. These concepts are demonstrated by revisiting two experiments from the literature in which the Cattaneo–Vernotte and the dual phase lag models are utilized. Our investigation revealed that these non-Fourier models are based on misinterpretations of the measured data, and the seeming deviation from Fourier’s law originates from the source terms and boundary conditions.

scholarly journals The impact of chromosomes and centrosomes on spindle assembly as observed in living cells.

1995 ◽  
Vol 129 (5) ◽  
pp. 1287-1300 ◽  
Author(s):  
D Zhang ◽  
R B Nicklas
Keyword(s):  
Spindle Assembly ◽  
Living Cells ◽  
Microtubule Dynamics ◽  
Specific Site ◽  
Polarization Microscopy ◽  
Spindle Microtubule ◽  
Single Chromosome ◽  
Bipolar Spindle ◽  
The Impact

We analyzed the role that chromosomes, kinetochores, and centrosomes play in spindle assembly in living grasshopper spermatocytes by reconstructing spindles lacking certain components. We used video-enhanced, polarization microscopy to distinguish the effect of each component on spindle microtubule dynamics and we discovered that both chromosomes and centrosomes make potent and very different contributions to the organization of the spindle. Remarkably, the position of a single chromosome can markedly affect the distribution of microtubules within a spindle or even alter the fate of spindle assembly. In an experimentally constructed spindle having only one chromosome, moving the chromosome to one of the two poles induces a dramatic assembly of microtubules at the nearer pole and a concomitant disassembly at the farther pole. So long as a spindle carries a single chromosome it will persist normally. A spindle will also persist even when all chromosomes are detached and then removed from the cell. If, however, a single chromosome remains in the cell but is detached from the spindle and kept in the cytoplasm, the spindle disassembles. One might expect the effect of chromosomes on spindle assembly to relate to a property of a specific site on each chromosome, perhaps the kinetochore. We have ruled out that possibility by showing that it is the size of chromosomes rather than the number of kinetochores that matters. Although chromosomes affect spindle assembly, they cannot organize a spindle in the absence of centrosomes. In contrast, centrosomes can organize a functional bipolar spindle in the absence of chromosomes. If both centrosomes and chromosomes are removed from the cell, the spindle quickly disappears.

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