The cell organization underlying structural colour is involved in Flavobacterium IR1 predation.

Flavobacterium IR1 is a gliding bacterium with a high degree of colonial organization as a 2D photonic crystal, resulting in vivid structural coloration when illuminated. Enterobacter cloacae B12, an unrelated bacterium, was isolated from the brown macroalga Fucus vesiculosus from the same location as IR1. IR1 was found to be a predator of B12. A process of surrounding, infiltration, undercutting and killing of B12 supported improved growth of IR1. A combination of motility and capillarity facilitated the engulfment of B12 colonies by IR1. Predation was independent of illumination. Mutants of IR1 that formed photonic crystals less effectively than the wild type were reduced in predation. Conversely, formation of a photonic crystal was not advantageous in resisting predation by Rhodococcus spp. PIR4. These observations suggest that the organization required to create structural colour has a biological function (facilitating predation) but one that is not directly related to the photonic properties of the colony. This work is the first experimental evidence supporting a role for this widespread type of cell organization in the Flavobacteriia.

Quantitative fluorescence imaging of protein diffusion and interaction in living cells.

Diffusion processes and local dynamic equilibria inside cells lead to nonuniform spatial distributions of molecules, which are essential for processes such as nuclear organization and signaling in cell division, differentiation and migration. To understand these mechanisms, spatially resolved quantitative measurements of protein abundance, mobilities and interactions are needed, but current methods have limited capabilities to study dynamic parameters. Here we describe a microscope based on light-sheet illumination that allows massively parallel fluorescence correlation spectroscopy (FCS) measurements and use it to visualize the diffusion and interactions of proteins in mammalian cells and in isolated fly tissue. Imaging the mobility of heterochromatin protein HP1α (ref. 4) in cell nuclei we could provide high-resolution diffusion maps that reveal euchromatin areas with heterochromatin-like HP1α-chromatin interactions. We expect that FCS imaging will become a useful method for the precise characterization of cellular reaction-diffusion processes.

Measuring and imaging diffusion with multiple scan speed image correlation spectroscopy.

The intracellular mobility of biomolecules is determined by transport and diffusion as well as molecular interactions and is crucial for many processes in living cells. Methods of fluorescence microscopy like confocal laser scanning microscopy (CLSM) can be used to characterize the intracellular distribution of fluorescently labeled biomolecules. Fluorescence correlation spectroscopy (FCS) is used to describe diffusion, transport and photo-physical processes quantitatively. As an alternative to FCS, spatially resolved measurements of mobilities can be implemented using a CLSM by utilizing the spatio-temporal information inscribed into the image by the scan process, referred to as raster image correlation spectroscopy (RICS). Here we present and discuss an extended approach, multiple scan speed image correlation spectroscopy (msICS), which benefits from the advantages of RICS, i.e. the use of widely available instrumentation and the extraction of spatially resolved mobility information, without the need of a priori knowledge of diffusion properties. In addition, msICS covers a broad dynamic range, generates correlation data comparable to FCS measurements, and allows to derive two-dimensional maps of diffusion coefficients. We show the applicability of msICS to fluorophores in solution and to free EGFP in living cells.

Multiscale analysis of the laser-induced damage threshold in optical coatings.

We have investigated the influence of laser beam size on laser-induced damage threshold (LIDT) in the case of single- and multiple-shot irradiation. The study was performed on hafnia thin films deposited with various technologies (evaporation, sputtering, with or without ion assistance). LIDT measurements were carried out at 1064 nm and 12 ns with a spot size ranging from a few tens to a few hundreds of micrometers, in 1-on-1 and R-on-1 modes. These measurements were compared with simulations obtained with the statistical theory of laser-induced damage caused by initiating inclusions. We show how to obtain information on the initiating defect properties and the related physical damage mechanisms with a multiscale study. Under certain conditions, it is possible with this method to discriminate different defects, estimate their densities, and follow the evolution of the defects under multiple irradiation. The different metrology implications of our approach, particularly for obtaining a functional LIDT of optical components are discussed.

Laser damage resistance of hafnia thin films deposited by electron beam deposition, reactive low voltage ion plating, and dual ion beam sputtering.

A comparative study is made of the laser damage resistance of hafnia coatings deposited on fused silica substrates with different technologies: electron beam deposition (from Hf or HfO(2) starting material), reactive low voltage ion plating, and dual ion beam sputtering. The laser damage thresholds of these coatings are determined at 1064 and 355 nm using a nanosecond pulsed YAG laser and a one-on-one test procedure. The results are associated with a complete characterization of the samples: refractive index n measured by spectrophotometry, extinction coefficient k measured by photothermal deflection, and roughness measured by atomic force microscopy.

Chalcogenide coatings of Ge15Sb20S65 and Te20As30Se50.

Chalcogenide coatings are investigated to obtain either optical components for spectral applications or optochemical sensors in the mid-infrared. The deposition of Ge(15)Sb(20)S(65) and Te(20)As(30)Se(50) chalcogenide glasses is performed by two physical techniques: electron-beam and pulsed-laser deposition. The quality of the film is analyzed by scanning electron microscopy, atomic force microscopy, and energy dispersive spectroscopy to characterize the morphology, topography, and chemical composition. The optical properties and optical constants are also determined. A CF(4) dry etching is performed on these films to obtain a channeled optical waveguide. For a passband filter made by electron-beam deposition, cryolite as a low-refractive-index material and chalcogenide glasses as high-refractive-index materials are used to favor a large refractive-index contrast. A shift of a centered wavelength of a photosensitive passband filter is controlled by illumination time.

Effect of laser irradiation on silica substrate contaminated by aluminum particles.

A major issue in the use of high-power lasers, such as the Laser Megajoule (LMJ), is laser-induced damage of optical components. One potential damage initiator is particulate contamination, but its effect is hard to distinguish from that of other damage precursors. To do so, we introduced artificial contaminants typical of metallic pollution likely to be present on the optical components of the LMJ chains. More precisely, aluminum particles of two different sizes were placed on a silica sample. These dots were characterized by optical microscopy and profilometry. Then they were exposed to a laser beam with a pulse length of 6.5 ns at 1064 nm and fluences in the range from 1 to 40 J/cm(2). Each dot was characterized again with the same techniques and also by photothermal microscopy. To complete the experimental results, we performed numerical simulations with a one-dimensional Lagrangian hydrodynamics code. We show that the particle removal by laser irradiation produces a modification of the silica surface that does not evolve into catastrophic damage under subsequent irradiation. However, the effect does depend on the size of the dots. We demonstrate that a procedure exists that removes the dot and leaves the site capable of resisting high fluence.

Enhancement of single-molecule fluorescence detection in subwavelength apertures.

We report the experimental proof of molecular count rate enhancement (up to 6.5-fold) and lifetime reduction for single fluorescent molecules diffusing in subwavelength apertures milled in aluminum films. The observed enhancement dependence with the aperture diameter agrees qualitatively with numerical electromagnetic computations of the excitation power density into the aperture volume.