313 research outputs found
Real-Time analysis and visualization for single-molecule based super-resolution microscopy
Accurate multidimensional localization of isolated fluorescent emitters is a time consuming process in single-molecule based super-resolution microscopy. We demonstrate a functional method for real-time reconstruction with automatic feedback control, without compromising the localization accuracy. Compatible with high frame rates of EM-CCD cameras, it relies on a wavelet segmentation algorithm, together with a mix of CPU/GPU implementation. A combination with Gaussian fitting allows direct access to 3D localization. Automatic feedback control ensures optimal molecule density throughout the acquisition process. With this method, we significantly improve the efficiency and feasibility of localization-based super-resolution microscopy
Quantum yield optimized fluorophores for site-specific labeling and super-resolution imaging
Single molecule applications, saturated pattern excitation microscopy, or
stimulated emission depletion (STED) microscopy demand for bright and highly
stable fluorescent dyes1,2. Despite of intensive research the choice of
fluorphores is still very limited. Typically a stable fluorescent dyes is
covalently attached to the target. This methodology brings forward a number of
limitations, in particular, in case of protein labeling. First of all the
fluorescent probes need to be attached selectively and site-specifically to
prevent unspecific background. This often requires single cysteine mutations
for covalent protein modification. Employing quantum dots allows overcoming
problems of photo-bleaching3-6. However, the downsides are their large size,
rendering the probe inaccessible to spatially confined architectures, issues in
biocompatibility due to proper particle coating, and cellular toxicity6-8. Here
we propose a new method to overcome the above outlined problems
A variational analysis of Einstein-scalar field Lichnerowicz equations on compact Riemannian manifolds
We establish new existence and non-existence results for positive solutions
of the Einstein-scalar field Lichnerowicz equation on compact manifolds. This
equation arises from the Hamiltonian constraint equation for the
Einstein-scalar field system in general relativity. Our analysis introduces
variational techniques, in the form of the mountain pass lemma, to the analysis
of the Hamiltonian constraint equation, which has been previously studied by
other methods.Comment: 15 page
Multiple Routes for Glutamate Receptor Trafficking: Surface Diffusion and Membrane Traffic Cooperate to Bring Receptors to Synapses
Trafficking of glutamate receptors into and out of synapses is critically
involved in the plasticity of excitatory synaptic transmission. Endocytosis and
exocytosis of receptors have initially been thought to account alone for this
trafficking. However, membrane proteins also traffic through surface lateral
diffusion in the plasma membrane. We describe developments in
electrophysiological and optical approaches that have allowed for the real time
measurement of glutamate receptor surface trafficking in live neurons. These
include (i) specific imaging of surface receptors using a pH sensitive
fluorescent protein, (ii) design of a photoactivable drug to inactivate locally
surface receptors and monitor electrophysiologically their recovery, and
(iii)application of single molecule fluorescence microscopy to directly track
the movement of individual surface receptors with nanometer resolution inside
and outside synapses. Altogether, these approaches have demonstrated that
glutamate receptors diffuse at high rates in the neuronal membrane and suggest
a key role for surface diffusion in the regulation of receptor numbers at
synapses
The constraint equations for the Einstein-scalar field system on compact manifolds
We study the constraint equations for the Einstein-scalar field system on
compact manifolds. Using the conformal method we reformulate these equations as
a determined system of nonlinear partial differential equations. By introducing
a new conformal invariant, which is sensitive to the presence of the initial
data for the scalar field, we are able to divide the set of free conformal data
into subclasses depending on the possible signs for the coefficients of terms
in the resulting Einstein-scalar field Lichnerowicz equation. For many of these
subclasses we determine whether or not a solution exists. In contrast to other
well studied field theories, there are certain cases, depending on the mean
curvature and the potential of the scalar field, for which we are unable to
resolve the question of existence of a solution. We consider this system in
such generality so as to include the vacuum constraint equations with an
arbitrary cosmological constant, the Yamabe equation and even (all cases of)
the prescribed scalar curvature problem as special cases.Comment: Minor changes, final version. To appear: Classical and Quantum
Gravit
Monitoring the emergence of antibiotic resistance using the technology ot the DebugIT platform in the HEGP context
International audiencepas de résum
A Septin-Dependent Diffusion Barrier at Dendritic Spine Necks
Excitatory glutamatergic synapses at dendritic spines exchange and modulate their receptor content via lateral membrane diffusion. Several studies have shown that the thin spine neck impedes the access of membrane and solute molecules to the spine head. However, it is unclear whether the spine neck geometry alone restricts access to dendritic spines or if a physical barrier to the diffusion of molecules exists. Here, we investigated whether a complex of septin cytoskeletal GTPases localized at the base of the spine neck regulates diffusion across the spine neck. We found that, during development, a marker of the septin complex, Septin7 (Sept7), becomes localized to the spine neck where it forms a stable structure underneath the plasma membrane. We show that diffusion of receptors and bulk membrane, but not cytoplasmic proteins, is slower in spines bearing Sept7 at their neck. Finally, when Sept7 expression was suppressed by RNA interference, membrane molecules explored larger membrane areas. Our findings indicate that Sept7 regulates membrane protein access to spines
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