Maxime Dahan

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Papers by Maxime Dahan

Fluorescence lifetime modification of single CdSe nanocrystals

Quantum Dots: Inorganic Fluorescent Probes for Single-Molecule Tracking Experiments in Live Cells

Des nanosources de lumieres pour l’optique et la biologie

$Research paper thumbnail of Single cell correlation fractal dimension of chromatin: a framework to interpret 3D single molecule super-resolution$

Single cell correlation fractal dimension of chromatin: a framework to interpret 3D single molecule super-resolution

10 d'Ulm Paris cedex 05 11 12 13 14 15 16 $) [email protected] 17 *) xavier.darzac... more 10 d'Ulm Paris cedex 05 11 12 13 14 15 16 $) [email protected] 17 *) [email protected] 18 19 No potential conflict of interest was disclosed 20 21 ) (r K distribution of H2B followed a power law, leading to a precise 33 measurement of the correlation fractal dimension of chromatin of 2.6. Moreover, using photoactivable 34 GFP linked to H2B, we measured the dynamics of different chromatin domains over several hours and 35 observed pronounced compaction and decompaction events. As a result, the correlation fractal 36 dimension of chromatin reported here can be interpreted as a measure of the distribution of sizes of 37 topologically segregated domains that constantly evolve with time. This highly dynamic view of 38 chromatin provides a solid framework to interpret the different levels of chromatin organization known 39 to regulate nuclear functions. 40 41 43 Récamier et al. (2013NUCLEUS0058) 3 44

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Single quantum dot tracking of membrane receptors

format_quoteQD labeling provides superior visualization of HER2 and EGFR signaling compared to traditional fluorescence methods.format_quote

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An atom trap relying on optical pumping

We have investigated a new radiation pressure trap which relies on optical pumping and does not r... more We have investigated a new radiation pressure trap which relies on optical pumping and does not require any magnetic field. I t employs six circularly polarized divergent beams and works on the red of a Jg + Je = Jg + 1 atomic transition with Jg 2 112. We have demonstrated this trap with cesium atoms from a vapour ce11 using the 852 nm Jg = 4 -+Je = 5 resonance transition. The trap contained up to 3.10' atoms in a cloud of 116 radius of 330 !lm.

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Imaging the lateral diffusion of membrane molecules with quantum dots

This protocol describes a sensitive approach to tracking the motion of membrane molecules such as... more This protocol describes a sensitive approach to tracking the motion of membrane molecules such as lipids and proteins with molecular resolution in live cells. This technique makes use of fluorescent semiconductor nanocrystals, quantum dots (QDs), as a probe to detect membrane molecules of interest. The photostability and brightness of QDs allow them to be tracked at a single particle level for longer periods than previous fluorophores, such as fluorescent proteins and organic dyes. QDs are bound to the extracellular part of the object to be followed, and their movements can be recorded with a fluorescence microscope equipped with a spectral lamp and a sensitive cooled charge-coupled device camera. The experimental procedure described for neurons below takes about 45 min. This technique is applicable to various cultured cells.

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Raman cooling of cesium below 3 nK: new approach inspired by Lévy flight statistics

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Single-molecule tracking in live cells reveals distinct target-search strategies of transcription factors in the nucleus

36 CNRS UMR 8197 37 46 rue d'Ulm 75230 Paris cedex 05 38 +33 (0) 1 44 32 34 62 39 [email protected] ... more 36 CNRS UMR 8197 37 46 rue d'Ulm 75230 Paris cedex 05 38 +33 (0) 1 44 32 34 62 39 [email protected] 40 2 Abstract 41 42 Gene regulation relies on transcription factors (TFs) exploring the nucleus 43 searching their targets. So far, most studies have focused on how fast TFs 44 diffuse, underestimating the role of nuclear architecture. We implemented a 45 single-molecule tracking assay to determine TFs dynamics. We found that c-46 Myc is a global explorer of the nucleus. In contrast, the positive transcription 47 elongation factor P-TEFb is a local explorer that oversamples its environment.

format_quotec-Myc and P-TEFb show different nuclear exploration strategies, emphasizing their unique functional roles in transcriptional control.format_quote

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Single quantum dot imaging in live cells: toward a cellular GPS

Colloidal semiconductor quantum dots (QDs) have become common fluorescent probes in biology. Thei... more Colloidal semiconductor quantum dots (QDs) have become common fluorescent probes in biology. Their optical properties not only facilitate spectrally multiplexed detection but also enable single molecule measurements with high signal to noise ratio. This is of particular interest in cell biology since it allows individual QD-tagged biomolecules to be tracked with good spatial and temporal resolution over long durations. Recent

Labeling Neuronal Membrane Receptors with Quantum Dots

Cold Spring Harbor Protocols, 2011

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Orientational imaging and tracking of single CdSe nanocrystals by defocused microscopy

Chemical Physics Letters, 2005

We study the fluorescence patterns produced by single CdSe nanocrystals in defocused microscopy. ... more We study the fluorescence patterns produced by single CdSe nanocrystals in defocused microscopy. The patterns are directly related to the structure of the nanocrystal emission dipole and their analysis provides a full determination of the three-dimensional orientation of the nanoparticles. In the case of a high numerical aperture objective, we show that the contour intensities of the defocused patterns can

Imaging of single molecules in live cells

M S-Medecine Sciences, 2011

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Wavelet analysis for single molecule localization microscopy

Optics Express, 2012

Localization of single molecules in microscopy images is a key step in quantitative single partic... more Localization of single molecules in microscopy images is a key step in quantitative single particle data analysis. Among them, single molecule based super-resolution optical microscopy techniques require high localization accuracy as well as computation of large data sets in the order of 10 5 single molecule detections to reconstruct a single image. We hereby present an algorithm based on image wavelet segmentation and single particle centroid determination, and compare its performance with the commonly used Gaussian fitting of the point spread function. We performed realistic simulations at different signal-to-noise ratios and particle densities and show that the calculation time using the wavelet approach can be more than one order of magnitude faster than that of Gaussian fitting without a significant degradation of the localization accuracy, from 1 nm to 4 nm in our range of study. We propose a simulation-based estimate of the resolution of an experimental single molecule acquisition.

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Single-molecule dynamics of enhanceosome assembly in embryonic stem cells

Cell, Jan 13, 2014

Enhancer-binding pluripotency regulators (Sox2 and Oct4) play a seminal role in embryonic stem (E... more Enhancer-binding pluripotency regulators (Sox2 and Oct4) play a seminal role in embryonic stem (ES) cell-specific gene regulation. Here, we combine in vivo and in vitro single-molecule imaging, transcription factor (TF) mutagenesis, and ChIP-exo mapping to determine how TFs dynamically search for and assemble on their cognate DNA target sites. We find that enhanceosome assembly is hierarchically ordered with kinetically favored Sox2 engaging the target DNA first, followed by assisted binding of Oct4. Sox2/Oct4 follow a trial-and-error sampling mechanism involving 84-97 events of 3D diffusion (3.3-3.7 s) interspersed with brief nonspecific collisions (0.75-0.9 s) before acquiring and dwelling at specific target DNA (12.0-14.6 s). Sox2 employs a 3D diffusion-dominated search mode facilitated by 1D sliding along open DNA to efficiently locate targets. Our findings also reveal fundamental aspects of gene and developmental regulation by fine-tuning TF dynamics and influence of the epigen...

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Magneto-fluorescent core-shell supernanoparticles

Nature Communications, 2014

Magneto-fluorescent particles have been recognized as an emerging class of materials that exhibit... more Magneto-fluorescent particles have been recognized as an emerging class of materials that exhibit great potential in advanced applications. However, synthesizing such magnetofluorescent nanomaterials that simultaneously exhibit uniform and tunable sizes, high magnetic content loading, maximized fluorophore coverage at the surface and a versatile surface functionality has proven challenging. Here we report a simple approach for co-assembling magnetic nanoparticles with fluorescent quantum dots to form colloidal magneto-fluorescent supernanoparticles. Importantly, these supernanoparticles exhibit a superstructure consisting of a close-packed magnetic nanoparticle 'core', which is fully surrounded by a 'shell' of fluorescent quantum dots. A thin layer of silica coating provides high colloidal stability and biocompatibility, and a versatile surface functionality. We demonstrate that after surface pegylation, these silica-coated magneto-fluorescent supernanoparticles can be magnetically manipulated inside living cells while being optically tracked. Moreover, our silica-coated magneto-fluorescent supernanoparticles can also serve as an in vivo multi-photon and magnetic resonance dual-modal imaging probe.

format_quoteEncapsulation of CS-SPs in silica significantly improves biocompatibility and colloidal stability for diverse applications.format_quote

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Fast multicolor 3D imaging using aberration-corrected multifocus microscopy

Nature methods, 2013

Conventional acquisition of three-dimensional (3D) microscopy data requires sequential z scanning... more Conventional acquisition of three-dimensional (3D) microscopy data requires sequential z scanning and is often too slow to capture biological events. We report an aberration-corrected multifocus microscopy method capable of producing an instant focal stack of nine 2D images. Appended to an epifluorescence microscope, the multifocus system enables high-resolution 3D imaging in multiple colors with single-molecule sensitivity, at speeds limited by the camera readout time of a single image.

$the nine focal planes; the CCG, with panels containing blazed diffractive gratings that reverse the dispersion of the MFG (central panel is blank); and the prism, which directs the images to their positions on the camera. Note the geometrical distortion of the MFG pattern, which introduces a phase shift the is dependent on diffractive order (m,, my) and gives rise to the focus shift in each plane of the multifocus image. (e) Raw, multifocus image of 200-nm fluorescent beads. As illustrated in c, the central tile is the (nondiffracted) traditional microscope image of the nominal focal plane. The surrounding eight tiles are the duplicate focus-shifted images, formed by diffraction in the MFG. The focus step between successive planes is Az = 380 nm. (f) Axial (xz) point-spread function (PSF), radially averaged and displayed in log scale. (g) Gaussian curves fitted to the bead signal of each plane at different Z positions of the stage. Best focus position is estimated as the maximum of the fitted curve (Supplementary Video 1). (h) Plot of the best focus position of each plane. The linear curve verifies the constant focus step Az between planes. Scale bars, 1 um.$

$Figure 2 | Multifocus imaging of yeast centromeres. (a,b) Multifocus transmission (a) and fluorescence (b) images of S. cerevisiae expressing Cse4-GFP. (c) Separation of the centromeres during anaphase over time (maximum-intensity projections). (d) Movement in 3D of the two centromere clusters (black and gray). Bottom right, separation between the centromere clusters over time. Rapid movement (phase I) is followed by slow movement (phase II). Inset, average spee during phases I and II (n = 5 cells). Scale bars, 5 um. to the theoretical maximum efficiency (~67%) of this type of grat- ing, but it could be improved (theoretically to ~93%) by using a multiphase element®. To implement an MFM microscope with more focal planes, one would use a different MFG to image the Mx My = 0, +1, £2 diffractive orders, obtaining 5 x 5 = 25 planes, as shown in Supplementary Figure 1.$

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Imaging the fate of histone Cse4 reveals de novo replacement in S phase and subsequent stable residence at centromeres

eLife, 2014

The budding yeast centromere contains Cse4, a specialized histone H3 variant. Fluorescence pulse-... more