Comparison between SOFI and STORM

A straightforward method to achieve super-resolution consists of taking an image sequence of stochastically blinking emitters using a standard wide-field fluorescence microscope. Densely packed single molecules can be distinguished sequentially in time using high-precision localization algorithms (e.g., PALM and STORM) or by analyzing the statistics of the temporal fluctuations (SOFI). In a face-to-face comparison of the two post-processing algorithms, we show that localization-based super-resolution can deliver higher resolution enhancements but imposes significant constraints on the blinking behavior of the probes, which limits its applicability for live-cell imaging. SOFI, on the other hand, works more consistently over different photo-switching kinetics and also delivers information about the specific blinking statistics. Its suitability for low SNR acquisition reveals SOFI's potential as a high-speed super-resolution imaging technique

Multiplane 3D superresolution optical fluctuation imaging

By switching fluorophores on and off in either a deterministic or a stochastic manner, superresolution microscopy has enabled the imaging of biological structures at resolutions well beyond the diffraction limit. Superresolution optical fluctuation imaging (SOFI) provides an elegant way of overcoming the diffraction limit in all three spatial dimensions by computing higher-order cumulants of image sequences of blinking fluorophores acquired with a conventional widefield microscope. So far, three-dimensional (3D) SOFI has only been demonstrated by sequential imaging of multiple depth positions. Here we introduce a versatile imaging scheme which allows for the simultaneous acquisition of multiple focal planes. Using 3D cross-cumulants, we show that the depth sampling can be increased. Consequently, the simultaneous acquisition of multiple focal planes reduces the acquisition time and hence the photo-bleaching of fluorescent markers. We demonstrate multiplane 3D SOFI by imaging the mitochondria network in fixed C2C12 cells over a total volume of $65\times65\times3.5 \mu\textrm{m}^3$ without depth scanning.Comment: 7 pages, 3 figure

Radar-based assessment of hail frequency in Europe

In this study we present a unique 10 year climatology of severe convective storm tracks for a large European area covering Germany, France, Belgium and Luxembourg. For the period 2005–2014, a high-resolution hail potential composite of 1×1 km$^{2}$ is produced from two-dimensional radar reflectivity and lightning data. Individual hailstorm tracks as well as their physical properties, such as radar reflectivity along the tracks, were reconstructed for the entire time period using the Convective Cell Tracking Algorithm (CCTA2D). A sea-to-continent gradient in the number of hail days per year is found to be present over the whole domain. In addition, the highest number of severe storms is found on the leeward side of low mountain ranges such as the Massif Central in France and the Swabian Jura in southwest Germany. A latitude shift in the hail peak month is observed between the northern part of Germany, where hail occurs most frequently in August, and southern France, where the maximum amount of hail is 2 months earlier. The longest footprints with high reflectivity values occurred on 9 June 2014 and on 28 July 2013 with lengths reaching up to 500 km. Both events were associated with hailstones measuring up to 10 cm diameter, which caused damage in excess of EUR 2 billion

Prototype de polarimètre micro-onde portable pour la détection à distance des températures de brillance

Ce mémoire de maîtrise décrit la première phase de la réalisation d'un prototype de polarimètre portable à 37GHz. Les polarimètres portables actuellement disponibles sur le marché coûtent au dessus de 200 000$ et sont plus adaptés à l'utilisation au laboratoire que sur le terrain. Ils nécessitent le transport de gaz à l'état liquide pour leur calibration et ont de grands besoins énergétiques. Tous ces facteurs contribuent à leur haut coût et à leur grande complexité d'utilisation. Cette première phase du projet vise à développer la partie analogique du récepteur micro-onde qui sera au coeur du prototype. D'autres travaux, à la suite de cette maîtrise, seront nécessaires afin de rendre le récepteur utilisable et compléter le prototype. Les spécifications du polarimètre ont d'abord été définies. Par la suite, la décision d'acheter ou de fabriquer chaque sous-système/circuit a été prise. Ensuite, les activités d'achat et de conception ont eu lieu. Il a été constaté que les techniques standards utilisées pour fabriquer des circuits microruban n'étaient pas assez précis. Un procédé de fabrication a alors été développé et les circuits ont été fabriqués. Le tout a finalement été intégré. La finalité de ce projet a été un système analogique qui permet d'amplifier et de traiter une onde incidente à haute fréquence et à faible intensité pour la rendre mesurable par un système d'échantillonnage et de traitement numérique. Ce système numérique sera conçu et fabriqué à une date ultérieure

How to display data by color schemes compatible with red-green color perception deficiencies

Visualization of data concerns most scientists. The use of color is required in order to display multidimensional information. In addition, color encoding a univariate image can improve the interpretation significantly. However up to 10% of the adult male population are affected by a red-green color perception deficiency which hampers the correct interpretation and appreciation of color encoded information. This work attempts to give guidelines on how to display a given dataset in a balanced manner. Three novel color maps are proposed providing readers with normal color perception a maximum of color contrast while being a good compromise for readers with color perception deficiencies. Please refer to http://lob.epfl.ch/page-89396.html for downloading these colormaps for your own usage

Feasiability of Using Evidence-Based Virtopsy to Answer the Possible Clinical and Post-Mortem Questions, in Veterinary Practice

A post-mortem examination is an important part of evidence-based medicine to understand the deterioration of clinical signs or causes of death in euthanized or deceased individual animals or even populations. Post-mortem analysis is aimed at improving clinical treatment and therapy, confirming a suspected diagnosis, man-aging breeding strategies, and clarifying the forensic cases (e.g., neglect or animal abuse). In analogy to virtopsy in human medicine, diagnostic imaging modalities have been applied in post-mortem veterinary medicine, which we call vetvirtopsy. We hypothesize that vetvirtopsy can be used as a method to answer certain clinical/post-mortem questions to im-prove the diagnosis reliability. In some questions, vetvirtopsy actually can replace conventional necropsy. This overview study aims to compare vetvirtopsy with conventional necropsy for variable causes of death in animals and to define its possibilities and limitations. Deceased or euthanized pets and wild animals were collected. The imaging techniques, such as post-mortem digital radiography, post-mortem ultrasound, post-mortem computed tomography, and post-mortem magnetic resonance tomography combined with image-guided tissue sampling, were used to address the open questions about clinical symptoms or causes of their death. The case series in this project showed that diagnostic imaging techniques are feasible in answering distinct ante-mortem and post-mortem clinical and forensic questions. However, there is an interdisciplinary collaboration between diagnostic imaging and sampling under imaging guidance

Spectral Cross-Cumulants for Multicolor Super-resolved SOFI Imaging

Super-resolution optical fluctuation imaging (SOFI) provides a resolution beyond the diffraction limit by analysing stochastic fluorescence fluctuations with higher-order statistics. Using nth order spatio-temporal cross-cumulants the spatial resolution as well as the sampling can be increased up to n-fold in all three spatial dimensions. In this study, we extend the cumulant analysis into the spectral domain and propose a novel multicolor super-resolution scheme. The simultaneous acquisition of two spectral channels followed by spectral cross-cumulant analysis and unmixing increase the spectral sampling. The number of discriminable fluorophore species is thus not limited to the number of physical detection channels. Using two color channels, we demonstrate spectral unmixing of three fluorophore species in simulations and multiple experiments with different cellular structures, fluorophores and filter sets. Based on an eigenvalue/ vector analysis we propose a scheme for an optimized spectral filter choice. Overall, our methodology provides a novel route for easy-to-implement multicolor sub-diffraction imaging using standard microscopes while conserving the spatial super-resolution property. This makes simultaneous multiplexed super-resolution fluorescence imaging widely accessible to the life science community interested to probe colocalization between two or more molecular species.Comment: main: 21 pages & 4 figures, supplementary 20 pages & 16 figure

Assessment of transferrin recycling by Triplet Lifetime Imaging in living cells

An optical method is presented that allows the measurement of the triplet lifetime of a fluorescent molecule. This is a characteristic specific to each fluorophore. Based on differences in triplet lifetimes of two fluorescent species (autofluorescence versus label), this novel approach measures relative quantities of a transmembrane receptor and associated fluorescently labeled ligand during its recycling in living cells. Similarly to fluorescence-lifetime based methods, our approach is almost insensitive to photobleaching. A simple theory for unmixing two known triplet lifetimes is presented along with validation of the method by measurements of transferrin recycling in a model system based on chinese hamster ovarian cells (CHO). Transferrin is the delivery carrier for Fe3+ to the cell

Fast focus field calculations

We present a method for fast calculation of the electromagnetic field near the focus of an objective with a high numerical aperture (NA). Instead of direct integration, the vectorial Debye diffraction integral is evaluated with the fast Fourier transform for calculating the electromagnetic field in the entire focal region. We generalize this concept with the chirp z transform for obtaining a flexible sampling grid and an additional gain in computation speed. Under the conditions for the validity of the Debye integral representation, our method yields the amplitude, phase and polarization of the focus field for an arbitrary paraxial input field in the aperture of the objective. Our fast calculation method is particularly useful for engineering the point-spread function or for fast image deconvolution. We present several case studies by calculating the focus fields of high NA oil immersion objectives for various amplitude, polarization and phase distributions of the input field. In addition, the calculation of an extended polychromatic focus field generated by a Bessel beam is presented. This extended focus field is of particular interest for Fourier domain optical coherence tomography because it preserves a lateral resolution of a few micrometers over an axial distance in the millimeter range