Browse
Submit Data
Databases
API
Help

Dataset Information

22 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Dynamic label-free imaging of lipid nanodomains.

ABSTRACT: Lipid rafts are submicron proteolipid domains thought to be responsible for membrane trafficking and signaling. Their small size and transient nature put an understanding of their dynamics beyond the reach of existing techniques, leading to much contention as to their exact role. Here, we exploit the differences in light scattering from lipid bilayer phases to achieve dynamic imaging of nanoscopic lipid domains without any labels. Using phase-separated droplet interface bilayers we resolve the diffusion of domains as small as 50 nm in radius and observe nanodomain formation, destruction, and dynamic coalescence with a domain lifetime of 220±60 ms. Domain dynamics on this timescale suggests an important role in modulating membrane protein function.

SUBMITTER: de Wit G

PROVIDER: S-EPMC4603517 | biostudies-literature | 2015 Oct

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

Dynamic label-free imaging of lipid nanodomains.

de Wit Gabrielle G Danial John S H JS Kukura Philipp P Wallace Mark I MI

Proceedings of the National Academy of Sciences of the United States of America 20150923 40

Lipid rafts are submicron proteolipid domains thought to be responsible for membrane trafficking and signaling. Their small size and transient nature put an understanding of their dynamics beyond the reach of existing techniques, leading to much contention as to their exact role. Here, we exploit the differences in light scattering from lipid bilayer phases to achieve dynamic imaging of nanoscopic lipid domains without any labels. Using phase-separated droplet interface bilayers we resolve the d ...[more]

PMID: 26401022

Similar Datasets

Direct label-free imaging of nanodomains in biomimetic and biological membranes by cryogenic electron microscopy.

Project description:The nanoscale organization of biological membranes into structurally and compositionally distinct lateral domains is believed to be central to membrane function. The nature of this organization has remained elusive due to a lack of methods to directly probe nanoscopic membrane features. We show here that cryogenic electron microscopy (cryo-EM) can be used to directly image coexisting nanoscopic domains in synthetic and bioderived membranes without extrinsic probes. Analyzing a series of single-component liposomes composed of synthetic lipids of varying chain lengths, we demonstrate that cryo-EM can distinguish bilayer thickness differences as small as 0.5 Å, comparable to the resolution of small-angle scattering methods. Simulated images from computational models reveal that features in cryo-EM images result from a complex interplay between the atomic distribution normal to the plane of the bilayer and imaging parameters. Simulations of phase-separated bilayers were used to predict two sources of contrast between coexisting ordered and disordered phases within a single liposome, namely differences in membrane thickness and molecular density. We observe both sources of contrast in biomimetic membranes composed of saturated lipids, unsaturated lipids, and cholesterol. When extended to isolated mammalian plasma membranes, cryo-EM reveals similar nanoscale lateral heterogeneities. The methods reported here for direct, probe-free imaging of nanodomains in unperturbed membranes open new avenues for investigation of nanoscopic membrane organization.

| S-EPMC7443941 | biostudies-literature

Label-free and charge-sensitive dynamic imaging of lipid membrane hydration on millisecond time scales.

Project description:Biological membranes are highly dynamic and complex lipid bilayers, responsible for the fate of living cells. To achieve this function, the hydrating environment is crucial. However, membrane imaging typically neglects water, focusing on the insertion of probes, resonant responses of lipids, or the hydrophobic core. Owing to a recent improvement of second-harmonic (SH) imaging throughput by three orders of magnitude, we show here that we can use SH microscopy to follow membrane hydration of freestanding lipid bilayers on millisecond time scales. Instead of using the UV/VIS resonant response of specific membrane-inserted fluorophores to record static SH images over time scales of >1,000 s, we SH imaged symmetric and asymmetric lipid membranes, while varying the ionic strength and pH of the adjacent solutions. We show that the nonresonant SH response of water molecules aligned by charge-dipole interactions with charged lipids can be used as a label-free probe of membrane structure and dynamics. Lipid domain diffusion is imaged label-free by means of the hydration of charged domains. The orientational ordering of water is used to construct electrostatic membrane potential maps. The average membrane potential depends quadratically on an applied external bias, which is modeled by nonlinear optical theory. Spatiotemporal fluctuations on the order of 100-mV changes in the membrane potential are seen. These changes imply that membranes are very dynamic, not only in their structure but also in their membrane potential landscape. This may have important consequences for membrane function, mechanical stability, and protein/pore distributions.

| S-EPMC5910843 | biostudies-literature

Label-free analysis of breast tissue polarity by Raman imaging of lipid phase.

Project description:The formation of the basoapical polarity axis in epithelia is critical for maintaining the homeostasis of differentiated tissues. Factors that influence cancer development notoriously affect tissue organization. Apical polarity appears as a specific tissue feature that, once disrupted, would facilitate the onset of mammary tumors. Thus, developing means to rapidly measure apical polarity alterations would greatly favor screening for factors that endanger the breast epithelium. A Raman scattering-based platform was used for label-free determination of apical polarity in live breast glandular structures (acini) produced in three-dimensional cell culture. The coherent anti-Stokes Raman scattering signal permitted the visualization of the apical and basal surfaces of an acinus. Raman microspectroscopy subsequently revealed that polarized acini lipids were more ordered at the apical membranes compared to basal membranes, and that an inverse situation occurred in acini that lost apical polarity upon treatment with Ca(2+)-chelator EGTA. This method overcame variation between different cultures by tracking the status of apical polarity longitudinally for the same acini. Therefore, the disruption of apical polarity by a dietary breast cancer risk factor, ?6 fatty acid, could be observed with this method, even when the effect was too moderate to permit a conclusive assessment by the traditional immunostaining method.

| S-EPMC3296051 | biostudies-literature

Three-dimensional label-free imaging and quantification of lipid droplets in live hepatocytes.

Project description:Lipid droplets (LDs) are subcellular organelles with important roles in lipid storage and metabolism and involved in various diseases including cancer, obesity, and diabetes. Conventional methods, however, have limited ability to provide quantitative information on individual LDs and have limited capability for three-dimensional (3-D) imaging of LDs in live cells especially for fast acquisition of 3-D dynamics. Here, we present an optical method based on 3-D quantitative phase imaging to measure the 3-D structural distribution and biochemical parameters (concentration and dry mass) of individual LDs in live cells without using exogenous labelling agents. The biochemical change of LDs under oleic acid treatment was quantitatively investigated, and 4-D tracking of the fast dynamics of LDs revealed the intracellular transport of LDs in live cells.

| S-EPMC5118789 | biostudies-literature

Label-free imaging of lipid depositions in C. elegans using third-harmonic generation microscopy.

Project description:Elucidation of the molecular mechanisms regulating lipid storage and metabolism is essential for mitigating excess adiposity and obesity, which has been associated with increased prevalence of severe pathological conditions such as cardiovascular disorders and type II diabetes, worldwide. However, imaging fatty acid distribution and dynamics in vivo, at the cellular or organismal level is challenging. We developed a label-free method for visualizing lipid depositions in vivo, based on third harmonic generation (THG) microscopy. THG imaging requires a single pulsed-laser light source, alleviating the technical challenges of implementing coherent anti-Stokes Raman scattering spectroscopy (CARS) to detect fat stores in living cells. We demonstrate that THG can be used to efficiently and reliably visualize lipid droplets in Caenorhabditis elegans. Thus, THG microscopy offers a versatile alternative to fluorescence and dye-based approaches for lipid biology research.

| S-EPMC3879321 | biostudies-literature

Dynamic nano-imaging of label-free living cells using electron beam excitation-assisted optical microscope.

Project description:Optical microscopes are effective tools for cellular function analysis because biological cells can be observed non-destructively and non-invasively in the living state in either water or atmosphere condition. Label-free optical imaging technique such as phase-contrast microscopy has been analysed many cellular functions, and it is essential technology for bioscience field. However, the diffraction limit of light makes it is difficult to image nano-structures in a label-free living cell, for example the endoplasmic reticulum, the Golgi body and the localization of proteins. Here we demonstrate the dynamic imaging of a label-free cell with high spatial resolution by using an electron beam excitation-assisted optical (EXA) microscope. We observed the dynamic movement of the nucleus and nano-scale granules in living cells with better than 100?nm spatial resolution and a signal-to-noise ratio (SNR) around 10. Our results contribute to the development of cellular function analysis and open up new bioscience applications.

| S-EPMC4630636 | biostudies-literature

Quantitative Label-Free Imaging of Lipid Domains in Single Bilayers by Hyperspectral Coherent Raman Scattering.

Project description:Lipid phase separation in cellular membranes is thought to play an important role in many biological functions. This has prompted the development of synthetic membranes to study lipid-lipid interactions in vitro, alongside optical microscopy techniques aimed at directly visualizing phase partitioning. In this context, there is a need to overcome the limitations of fluorescence microscopy, where added fluorophores can significantly perturb lipid packing. Raman-based optical imaging is a promising analytical tool for label-free chemically specific microscopy of lipid bilayers. In this work, we demonstrate the application of hyperspectral coherent Raman scattering microscopy combined with a quantitative unsupervised data analysis methodology developed in-house to visualize lipid partitioning in single planar membrane bilayers exhibiting liquid-ordered and liquid-disordered domains. Two home-built instruments were utilized, featuring coherent anti-Stokes Raman scattering and stimulated Raman scattering modalities. Ternary mixtures of dioleoylphosphatidylcholine, sphingomyelin, and cholesterol were used to form phase-separated domains. We show that domains are consistently resolved, both chemically and spatially, in a completely label-free manner. Quantitative Raman susceptibility spectra of the domains are provided alongside their spatially resolved concentration maps.

| S-EPMC7660592 | biostudies-literature

Shortwave-infrared meso-patterned imaging enables label-free mapping of tissue water and lipid content.

Project description:Water and lipids are key participants in many biological processes, but there are few non-invasive methods that provide quantification of these components in vivo, and none that can isolate and quantify lipids in the blood. Here we develop a new imaging modality termed shortwave infrared meso-patterned imaging (SWIR-MPI) to provide label-free, non-contact, spatial mapping of water and lipid concentrations in tissue. The method utilizes patterned hyperspectral illumination to target chromophore absorption bands in the 900-1,300 nm wavelength range. We use SWIR-MPI to monitor clinically important physiological processes including edema, inflammation, and tumor lipid heterogeneity in preclinical models. We also show that SWIR-MPI can spatially map blood-lipids in humans, representing an example of non-invasive and contact-free measurements of in vivo blood lipids. Together, these results highlight the potential of SWIR-MPI to enable new capabilities in fundamental studies and clinical monitoring of major conditions including obesity, cancer, and cardiovascular disease.

| S-EPMC7585425 | biostudies-literature

Label-free third harmonic generation imaging and quantification of lipid droplets in live filamentous fungi.

Project description:We report the utilization of Third-Harmonic Generation microscopy for label-free live cell imaging of lipid droplets in the hypha of filamentous fungus Phycomyces blakesleeanus. THG microscopy images showed bright spherical features dispersed throughout the hypha cytoplasm in control conditions and a transient increase in the number of bright features after complete nitrogen starvation. Colocalization analysis of THG and lipid-counterstained images disclosed that the cytoplasmic particles were lipid droplets. Particle Size Analysis and Image Correlation Spectroscopy were used to quantify the number density and size of lipid droplets. The two analysis methods both revealed an increase from 16 × 10-3 to 23 × 10-3 lipid droplets/µm2 after nitrogen starvation and a decrease in the average size of the droplets (range: 0.5-0.8 µm diameter). In conclusion, THG imaging, followed by PSA and ICS, can be reliably used for filamentous fungi for the in vivo quantification of lipid droplets without the need for labeling and/or fixation. In addition, it has been demonstrated that ICS is suitable for THG microscopy.

| S-EPMC9637149 | biostudies-literature

Lipid nanodomains change ion channel function.

Project description:Signaling proteins and neurotransmitter receptors often associate with saturated chain and cholesterol-rich domains of cell membranes, also known as lipid rafts. The saturated chains and high cholesterol environment in lipid rafts can modulate protein function, but evidence for such modulation of ion channel function in lipid rafts is lacking. Here, using raft-forming model membrane systems containing cholesterol, we show that lipid lateral phase separation at the nanoscale level directly affects the dissociation kinetics of the gramicidin dimer, a model ion channel.

| S-EPMC5599369 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data