Search results for "Fluorescence Correlation Spectroscopy"
showing 10 items of 29 documents
FRET-based dynamic structural biology: Challenges, perspectives and an appeal for open-science practices.
2021
International audience; Single-molecule FRET (smFRET) has become a mainstream technique for studying biomolecular structural dynamics. The rapid and wide adoption of smFRET experiments by an ever- increasing number of groups has generated significant progress in sample preparation, measurement procedures, data analysis, algorithms and documentation. Several labs that employ smFRET approaches have joined forces to inform the smFRET community about streamlining how to perform experiments and analyze results for obtaining quantitative information on biomolecular structure and dynamics. The recent efforts include blind tests to assess the accuracy and the precision of smFRET experiments among d…
Modeling of Particle Number Fluctuations in Entire Cells
2012
In a recent study we developed a method to model protein diffusion in cells [1], where special attention was given to generating from image data of the measured cell a realistic digital model cell in which protein dynamics were simulated. The method was shown to be well suited for modeling non-equilibrium situations that arise, e.g., in photobleaching experiments, and to be capable of producing more detailed information about protein motion than traditional modeling.Another experimental way to assess protein dynamics is to study fluctuations in the local protein number, as it is done, e.g., in fluorescence correlation spectroscopy (FCS), or in similar measurements that apply single-plane il…
Modifying the body distribution of HPMA-based copolymers by molecular weight and aggregate formation.
2011
There is a recognized need to create well-defined polymer probes for in vivo and clinical positron emission tomography (PET) imaging to guide the development of new generation polymer therapeutics. Using the RAFT polymerization technique in combination with the reactive ester approach, here we have synthesized well-defined and narrowly distributed N-(2-hydroxypropyl)methacrylamide homopolymers (pHPMA) (P1* and P2*) and random HPMA copolymers consisting of hydrophilic HPMA and hydrophobic lauryl methacrylate comonomers (P3* and P4*). The polymers had molecular weights below (P1* and P3*) and above the renal threshold (P2* and P4*). Whereas the homopolymers dissolve in isotonic solution as in…
Dye-Labeled Poly(organosiloxane) Microgels with Core−Shell Architecture
1999
Poly(organosiloxane) microgels are highly cross-linked rather monodisperse spherical particles of radius about 10 nm. Using a functionalized silane comonomer, i.e., (chlorobenzyl)trimethoxysilane, model particles suitable for studies in colloid physics are available: photoreactive and fluorescent dyes can be covalently bound within the microgels to prepare tracers for diffusion studies using forced Rayleigh scattering (FRS) and fluorescence correlation spectroscopy (FCS). For the application as tracer particles, it is important not to influence the diffusion behavior by the coupled chromophores. Therefore, functionalized precursors with a core−shell architecture are used to minimize labeli…
Fluorescence Decay Time of Single Semiconductor Nanocrystals
2002
We present fluorescence decay measurements of single ZnS covered CdSe nanocrystals. It is shown that the fluorescence decay time is fluctuating during the investigation leading to a multiexponential decay even for a single nanocrystal. In combination with measurements of the fluorescence blinking behavior we find that a high fluorescence intensity is correlated with a long fluorescence decay time. This is consistent with a model of fluctuating nonradiative decay channels leading to variable dynamic quenching processes of the excited state.
Protein diffusion in mammalian cell cytoplasm.
2011
We introduce a new method for mesoscopic modeling of protein diffusion in an entire cell. This method is based on the construction of a three-dimensional digital model cell from confocal microscopy data. The model cell is segmented into the cytoplasm, nucleus, plasma membrane, and nuclear envelope, in which environment protein motion is modeled by fully numerical mesoscopic methods. Finer cellular structures that cannot be resolved with the imaging technique, which significantly affect protein motion, are accounted for in this method by assigning an effective, position-dependent porosity to the cell. This porosity can also be determined by confocal microscopy using the equilibrium distribut…
Disassembly of structurally modified viral nanoparticles: characterization by fluorescence correlation spectroscopy.
2005
Abstract Analysis of the breakdown products of engineered viral particles can give useful information on the particle structure. We used various methods to breakdown both a recombinant enveloped virus and virus-like particles (VLPs) from two non-enveloped viruses and analysed the resulting subunits by fluorescence correlation spectroscopy (FCS). Analysis of the enveloped baculovirus, Autographa californica multicapsid nucleopolyhedrovirus (AcMNPV), displaying the green fluorescent protein (GFP) fused to its envelope protein gp64 was performed in the presence and absence of 5 mM SDS and 25 mM DTT. Without treatment, the viral particle showed a diffusion time of 3.3 ms. In the presence of SDS…
Scanning near-field optical microscopy using semiconductor nanocrystals as a local fluorescence and fluorescence resonance energy transfer source
2003
Local fluorescence probes based on CdSe semiconductor nanocrystals were prepared and tested by recording scanning near-field optical microscopy (SNOM) images of calibration samples and fluorescence resonance energy transfer SNOM (FRET SNOM) images of acceptor dye molecules inhomogeneously deposited onto a glass substrate. Thousands of nanocrystals contribute to the signal when this probe is used as a local fluorescence source while only tens of those (the most apical) are involved in imaging for the FRET SNOM operation mode. The dip-coating method used to make the probe enables diminishing the number of active fluorescent nanocrystals easily. Prospects to realize FRET SNOM based on a single…
Monitoring human parvovirus B19 virus-like particles and antibody complexes in solution by fluorescence correlation spectroscopy
2004
AbstractFluorescence correlation spectroscopy (FCS) was used in monitoring human parvovirus B19 virus-like particle (VLP) antibody complexes from acute phase and pastimmunity serum samples. The Oregon Green 488-labeled VLPs gave an average diffusion coefficient of 1.7x10exp-7 cm(2)s(-1) with an apparent hydrodynamic radius of 14 nm. After incubation of the fluorescent VLPs with an acute phase serum sample, the mobility information obtained from the fluorescence intensity fluctuation by autocorrelation analysis showed an average diffusion coefficient of 1.5x10exp-8 cm(2)s(-1), corresponding to an average radius of 157 nm. In contrast, incubation of the fluorescent VLPs with a pastimmunity se…
Time resolved confocal luminescence investigations on reverse proton exchange Nd : LiNbO3 channel waveguides
2009
In this work we report on the time and spatial resolved fluorescence of Neodymium ions in LiNbO(3) channel waveguides fabricated by Reverse Proton Exchange. The analysis of the fluorescence decay curves obtained with a sub-micrometric resolution has evidenced the presence of a relevant fluorescence quenching inside the channel waveguide. From the comparison between diffusion simulations and the spatial dependence of the (4)F(3/2) fluorescence decay rate we have concluded that the observed fluorescence quenching can be unequivocally related to the presence of H+ ions in the LiNbO(3) lattice. Nevertheless, it turns out that Reverse Proton Exchange guarantees a fluorescence quenching level sig…