A molecular rotor for measuring viscosity in plasma membranes of live cells

2014 ◽  
Vol 50 (40) ◽  
pp. 5282-5284 ◽  
Author(s):  
Ismael López-Duarte ◽  
Thanh Truc Vu ◽  
M. Angeles Izquierdo ◽  
James A. Bull ◽  
Marina K. Kuimova
Keyword(s):  
Plasma Membrane ◽  
Plasma Membranes ◽  
Live Cells ◽  
Molecular Rotor

We report the viscosity imaging of the plasma membrane of live cells using a new targeted BODIPY-based molecular rotor.

Imaging plasma membrane phase behaviour in live cells using a thiophene-based molecular rotor

2016 ◽  
Vol 52 (90) ◽  
pp. 13269-13272 ◽  
Author(s):  
Michael R. Dent ◽  
Ismael López-Duarte ◽  
Callum J. Dickson ◽  
Phoom Chairatana ◽  
Harry L. Anderson ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Lipid Bilayers ◽  
Plasma Membranes ◽  
Live Cell ◽  
Phase Behaviour ◽  
Live Cells ◽  
Molecular Rotor ◽  
Membrane Phase ◽  
Artificial Lipid Bilayers ◽  
Cell Plasma

A thiophene-based molecular rotor was used to probe ordering and viscosity within artificial lipid bilayers and live cell plasma membranes.

scholarly journals Imaging FCS Delineates Subtle Heterogeneity in Plasma Membranes of Resting Mast Cells

2019 ◽  
Author(s):  
Nirmalya Bag ◽  
David A. Holowka ◽  
Barbara A. Baird
Keyword(s):  
Plasma Membrane ◽  
Steady State ◽  
Mast Cells ◽  
Actin Polymerization ◽  
Plasma Membranes ◽  
Correlation Spectroscopy ◽  
Live Cells ◽  
Resting Cells ◽  
Functional Protein ◽  
Diffusion Properties

ABSTRACTA myriad of transient, nanoscopic lipid- and protein-based interactions confer a steady-state organization of plasma membrane in resting cells that is poised to orchestrate assembly of key signaling components upon reception of an extracellular stimulus. Although difficult to observe directly in live cells, these subtle interactions can be discerned by their impact on the diffusion of membrane constituents. Herein, we quantified the diffusion properties of a panel of structurally distinct lipid-anchored and transmembrane (TM) probes in RBL mast cells by multiplexed Imaging Fluorescence Correlation Spectroscopy. We developed a statistical analysis of data combined from many pixels over multiple cells to characterize differences as small as 10% in diffusion coefficients, which reflect differences in underlying interactions. We found that the distinctive diffusion properties of lipid-anchored probes can be explained by their dynamic partitioning into ordered proteo-lipid nanodomains, which encompass a major fraction of the membrane and whose physical properties are influenced by actin polymerization. Effects on diffusion by functional protein modules in both lipid-anchored and TM probes reflect additional complexity in steady-state membrane organization. The contrast we observe between different probes diffusing through the same membrane milieu represent the dynamic resting steady-state, which serves as a baseline for monitoring plasma membrane remodeling that occurs upon stimulation.

scholarly journals Nanoscale spatio-temporal diffusion modes measured by simultaneous confocal and STED imaging

2018 ◽  
Author(s):  
Falk Schneider ◽  
Dominic Waithe ◽  
Silvia Galiani ◽  
Jorge Bernadino de la Serna ◽  
Erdinc Sezgin ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Plasma Membranes ◽  
Molecular Diffusion ◽  
Correlation Spectroscopy ◽  
Stimulated Emission ◽  
Temporal Organization ◽  
Live Cells ◽  
Apparent Diffusion Coefficients ◽  
Diffusion Dynamics ◽  
Spatio Temporal

AbstractThe diffusion dynamics in the cellular plasma membrane provides crucial insights into the molecular interactions, organization and bioactivity. Fluorescence correlation spectroscopy combined with super-resolution stimulated emission depletion nanoscopy (STED-FCS) measures such dynamics with high spatial and temporal resolution and reveals nanoscale diffusion characteristics by measuring the molecular diffusion in conventional confocal mode and super-resolved STED mode sequentially. However, to directly link the spatial and the temporal information, a method that simultaneously measures the diffusion in confocal and STED modes is needed. Here, to overcome this problem, we establish an advanced STED-FCS measurement method; line interleaved excitation scanning STED-FCS (LIESS-FCS) which discloses the molecular diffusion modes at different spatial positions with a single measurement. It relies on fast beam-scanning along a line with alternating laser illumination that yields, for each pixel, the apparent diffusion coefficients for two different observation spot sizes (conventional confocal and super-resolved STED). We demonstrate the potential of the LIESS-FCS approach with simulations and experiments on lipid diffusion in model and live cell plasma membranes. We also apply LIESS-FCS to investigate the spatio-temporal organization of GPI-anchored proteins in the plasma membrane of live cells which interestingly show multiple diffusion modes at different spatial positions.

Targetable, two-photon fluorescent probes for local nitric oxide capture in the plasma membranes of live cells and brain tissues

The Analyst ◽  
2018 ◽  
Vol 143 (17) ◽  
pp. 4180-4188 ◽  
Author(s):  
Xinfu Zhang ◽  
Benlei Wang ◽  
Yi Xiao ◽  
Chao Wang ◽  
Ling He
Keyword(s):  
Nitric Oxide ◽  
Plasma Membrane ◽  
Fluorescent Probe ◽  
Fluorescent Probes ◽  
Plasma Membranes ◽  
Live Cells ◽  
Two Photon ◽  
In Cells ◽  
Brain Tissues

A plasma membrane-targetable two-photon fluorescent probe for capturing nitric oxide in cells and brain tissues.

scholarly journals Isolation of plasma and nuclear membranes of thymocytes. II. Biochemical composition

1978 ◽  
Vol 77 (1) ◽  
pp. 232-245 ◽  
Author(s):  
A Monneron ◽  
J d'Alayer
Keyword(s):  
Plasma Membrane ◽  
Sialic Acid ◽  
Biochemical Composition ◽  
Plasma Membranes ◽  
Specific Activity ◽  
Sodium Dodecyl ◽  
High Specific Activity ◽  
Live Cells ◽  
Protein Ratio ◽  
Nuclear Membranes

Thymocyte plasma and nuclear membranes obtained by the procedure described in the accompanying paper were analyzed for their biochemical composition. Plasma membranes were very rich in phospholipid, cholesterol, sialic aicd; they did not contain nucleic acids. In comparison, nuclear membranes had a lower phospholipid to protein ratio and contained much less sialic acid and cholesterol. 50% of the cellular cholesterol and of the membrane-bound sialic acid were found in the plasma membranes, 14% in the nuclear membranes. Live cells were labeled with 131I, and the acid-insoluble radioactivity was followed in the subfractions. A good correlation with the distribution and enrichment of plasma membrane market-enzymes was obtained. Label enrichment was about 50-fold in the two lightest of the three plasma membrane fractions. 60% of the label was contained in the plasma membranes, only 4% in the nuclear membranes. Cross-contamination of these two types of membranes was thus negligible. Sodium dodecyl sulfate-gel electrophoresis revealed three different patterns specific for, respectively, plasma membranes, the microsomal-mitochondrial fraction, and nuclear membranes. Each pattern was characterized by a set of proteins and glycoproteins, among which high molecular weight glycoproteins could be considered as marker-proteins of, respectively, 280,000, 260,000, and 230,000 daltons. 131I-labeling of live cells tagged with a very high specific activity three glycoproteins of mol wt 280,000, 200,000, and 135,000 daltons. Nuclear membranes prepared from labeled isolated nuclei had a set of labeled proteins completely different from plasma membranes.

Membrane microdomains: lessons from microscopy

1995 ◽  
Vol 53 ◽  
pp. 776-777
Author(s):  
J.M. Robinson ◽  
J.M Oliver
Keyword(s):  
Spatial Distribution ◽  
Plasma Membrane ◽  
Epithelial Cells ◽  
Plasma Membranes ◽  
Cell Types ◽  
Imaging Modalities ◽  
Membrane Microdomains ◽  
Membrane Domains ◽  
Lateral Heterogeneity ◽  
Functional Importance

Specialized regions of plasma membranes displaying lateral heterogeneity are the focus of this Symposium. Specialized membrane domains are known for certain cell types such as differentiated epithelial cells where lateral heterogeneity in lipids and proteins exists between the apical and basolateral portions of the plasma membrane. Lateral heterogeneity and the presence of microdomains in membranes that are uniform in appearance have been more difficult to establish. Nonetheless a number of studies have provided evidence for membrane microdomains and indicated a functional importance for these structures.This symposium will focus on the use of various imaging modalities and related approaches to define membrane microdomains in a number of cell types. The importance of existing as well as emerging imaging technologies for use in the elucidation of membrane microdomains will be highlighted. The organization of membrane microdomains in terms of dimensions and spatial distribution is of considerable interest and will be addressed in this Symposium.

Pore-Spanning Plasma Membranes Derived from Giant Plasma Membrane Vesicles

2021 ◽  
Author(s):  
Nikolas K. Teiwes ◽  
Ingo Mey ◽  
Phila C. Baumann ◽  
Lena Strieker ◽  
Ulla Unkelbach ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Plasma Membranes ◽  
Membrane Vesicles ◽  
Plasma Membrane Vesicles

scholarly journals Resolution of diaminobenzidine for the detection of horseradish peroxidase on surfaces of cultured cells.

1985 ◽  
Vol 33 (8) ◽  
pp. 837-839 ◽  
Author(s):  
A Messing ◽  
A Stieber ◽  
N K Gonatas
Keyword(s):  
Plasma Membrane ◽  
Horseradish Peroxidase ◽  
Plasma Membranes ◽  
Cultured Cells ◽  
Lateral Spread ◽  
Ciliary Ganglion ◽  
Monolayer Cultures ◽  
Indirect Immunoperoxidase ◽  
Ciliary Ganglion Neurons ◽  
Ganglion Neurons

The resolution of indirect immunoperoxidase methods for localizing antigens on the surface of plasma membranes of cultured cells was tested using dissociated monolayer cultures of ciliary ganglion neurons prelabeled with cationic ferritin. Clusters of ferritin were produced on the cell surface by warming the cells to 37 degrees C after the ferritin, rabbit anti-ferritin, and goat anti-rabbit immunoglobulin coupled to horseradish peroxidase had all been applied. Intense 3,3'-diaminobenzidine tetrahydrochloride (DAB) staining was limited to the regions immediately surrounding the ferritin clusters. The lateral spread of the DAB reaction product beyond the outer ferritin particles in each cluster averaged 54-81 nm in four experiments. A second type of increased density, coinciding with the thickness of the plasma membrane, was also seen. These stained plasma membranes extended 161-339 nm from the ferritin clusters.

scholarly journals Charged anaesthetics alter LM-fibroblast plasma-membrane enzymes by selective fluidization of inner or outer membrane leaflets

1986 ◽  
Vol 239 (2) ◽  
pp. 301-310 ◽  
Author(s):  
W D Sweet ◽  
F Schroeder
Keyword(s):  
Plasma Membrane ◽  
Active Site ◽  
Plasma Membranes ◽  
Local Anaesthetics ◽  
Cationic Drugs ◽  
Cell Plasma ◽  
Composition And Structure ◽  
Highly Sensitive ◽  
Functional Consequences ◽  
Anionic Drugs

The functional consequences of the differences in lipid composition and structure between the two leaflets of the plasma membrane were investigated. Fluorescence of 1,6-diphenylhexa-1,3,5-triene(DPH), quenching, and differential polarized phase fluorimetry demonstrated selective fluidization by local anaesthetics of individual leaflets in isolated LM-cell plasma membranes. As measured by decreased limiting anisotropy of DPH fluorescence, cationic (prilocaine) and anionic (phenobarbital and pentobarbital) amphipaths preferentially fluidized the cytofacial and exofacial leaflets respectively. Unlike prilocaine, procaine, also a cation, fluidized both leaflets of these membranes equally. Pentobarbital stimulated 5′-nucleotidase between 0.1 and 5 mM and inhibited at higher concentrations, whereas phenobarbital only inhibited, at higher concentrations. Cationic drugs were ineffective. Two maxima of (Na+ + K+)-ATPase activation were obtained with both anionic drugs. Only one activation maximum was obtained with both cationic drugs. The maximum in activity below 1 mM for all four drugs clustered about a single limiting anisotropy value in the cytofacial leaflet, whereas there was no correlation between activity and limiting anisotropy in the exofacial leaflets. Therefore, although phenobarbital and pentobarbital below 1 mM fluidized the exofacial leaflet more than the cytofacial leaflet, the smaller fluidization in the cytofacial leaflet was functionally significant for (Na+ + K+)-ATPase. Mg2+-ATPase was stimulated at 1 mM-phenobarbital, unaffected by pentobarbital and slightly stimulated by both cationic drugs at concentrations fluidizing both leaflets. Thus the activity of (Na+ + K+)-ATPase was highly sensitive to selective fluidization of the leaflet containing its active site, whereas the other enzymes examined were little affected by fluidization of either leaflet.

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