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Synchrotrons and modern FEL light sources operate with bunch repetition rates in the MHz range. The profile of the electron beam inside the accelerator can be characterized with indirect experimental techniques where linear array detectors are employed to measure the emitted synchrotron radiation or the spectrum of a near-IR laser. To improve the performance of modern beam diagnostics we have developed KALYPSO, a detector system operating with a continuous frame rate of up to 2.7 MHz. To facilitate the integration in different experiments, a modular architecture has been adopted. Different semiconductor micro-strip sensors can be connected to front-end ASICs to optimize the quantum efficiency at different photon energies, ranging from visible light up to near-IR. The front-end electronics are integrated within an heterogeneous DAQ consisting of FPGAs and GPUs, which allows scientists to implement real-time data processing algorithms. The current version of the detector is in operation at the KARA synchrotron light source and at the European XFEL. In this contribution we present the detector architecture, the performance results and the on-going technical developments.

Algae are the main primary producers in aquatic environments and therefore of fundamental importance for the global ecosystem. Mid-infrared (IR) microspectroscopy is a non-invasive tool that allows in principle studying chemical composition on a single-cell level. For a long time, however, mid-infrared (IR) imaging of living algal cells in an aqueous environment has been a challenge due to the strong IR absorption of water. In this study, we employed multi-beam synchrotron radiation to measure time-resolved IR hyperspectral images of individual Thalassiosira weissflogii cells in water in the course of acclimation to an abrupt change of CO2 availability (from 390 to 5000 ppm and vice versa) over 75 min. We used a previously developed algorithm to correct sinusoidal interference fringes from IR hyperspectral imaging data. After preprocessing and fringe correction of the hyperspectral data, principal component analysis (PCA) was performed to assess the spatial distribution of organic pools within the algal cells. Through the analysis of 200,000 spectra, we were able to identify compositional modifications associated with CO2 treatment. PCA revealed changes in the carbohydrate pool (1200–950 cm$$^{-1}$$ - 1 ), lipids (1740, 2852, 2922 cm$$^{-1}$$ - 1 ), and nucleic acid (1160 and 1201 cm$$^{-1}$$ - 1 ) as the major response of exposure to elevated CO2 concentrations. Our results show a local metabolism response to this external perturbation.

A bottleneck for the investigation of electron beam dynamics in ring accelerators is a fast detection scheme coping with their high repetition rates in the MHz range. For example, at KARA (KArlsruhe Research Accelerator), the electron storage ring at the Karlsruhe Institute of Technology (KIT) in Germany, we showed that electro-optical methods enable single-shot detection of longitudinal electron bunch profiles by imprinting them onto chirped laser pulses. However, commercial cameras required to detect the spectra are typically limited to hundreds of kHz in readout speed. To tackle these challenges, we developed KALYPSO (KArlsruhe Linear array detector for MHz-rePetition rate SpectrOscopy), a linear detector array with a data acquisition system (DAQ) allowing high data-rates over long time scales. Due to a modular approach, various sensors (InGaAs and Si) can be used, so that KALYPSO can be adapted to different experiments with spectral regimes ranging from near-ultraviolet (NUV) to near-infrared (NIR). In this talk, we present recent results on studies of longitudinal and horizontal bunch profiles using KALYPSO. As an outlook, we give another example for MHz-range readout using KALYPSO, namely horizontal bunch profile diagnostics measuring the radiation emitted from a dispersive section in a storage ring. At the KARA visible light diagnostics (VLD) port the emitted radiation above 400 nm was previously recorded with a fast-gated camera. Here, limitations are the repetition rate in combination with the huge number of cycles during continuous measurements. To overcome these limitations, KALYPSO can repleace the fast-gated camera. Furthermore, due to the easy implementation of KALYPSO, we envision numerous applications for table-top experiments as well as for large-scale facilies.

KALYPSO is a novel detector operating at line rates above 10 Mfps. It consists of a detector board connected to FPGA based readout card for real time data processing. The detector board holds a Si or InGaAs linear array sensor, with spectral sensitivity ranging from 400 nm to 2600 nm, which is connected to a custom made front-end ASIC. A FPGA readout framework performs the real time data processing. In this contribution, we present the detector system, the readout electronics and the heterogeneous infrastructure for machine learning processing. The detector is currently in use at several synchrotron facilities for beam diagnostics as well as for single-pulse laser characterizations. Thanks to the shot-to-shot capability over long time scale, new attractive applications are open up for imaging in biological and medical research.

The development of fast detection methods for comprehensive monitoring of electron bunches is a prerequisite to gain comprehensive control over the synchrotron emission in storage rings with their MHz repetition rate. Here, we present a proof-of-principle experiment with a detailed description of our implementation to detect the longitudinal electron bunch profiles via single-shot, near-field electro-optical sampling at the Karlsruhe Research Accelerator (KARA). Our experiment is equipped with an ultrafast line array camera providing a high-throughput MHz data stream. We characterize statistical properties of the obtained data set and give a detailed description for the data processing as well as for the calculation of the charge density profiles, which were measured in the short-bunch operation mode of KARA. Finally, we discuss properties of the bunch profile dynamics on a coarse-grained level for the example of the well-known synchrotron oscillation.

KALYPSO is a novel detector operating at line rates above 10 Mfps. The detector board holds a silicon or InGaAs linear array sensor with spectral sensitivity ranging from 400 nm to 2600 nm. The sensor is connected to a cutting-edge, custom designed, ASIC readout chip, which is responsible for the remarkable frame rate. The FPGA readout architecture enables continuous data acquisition and processing in real time. This detector is currently employed in many synchrotron facilities for beam diagnostics and for the characterization of self-built Ytterbium-doped fiber laser emitting around 1050 nm with a bandwidth of 40 nm.

To understand and control the dynamics in the longitudinal phase space, time-resolved measurements of different bunch parameters are required. For a reconstruction of this phase space, the detector systems have to be synchronized. This reconstruction can be used e.g. for studies of the micro-bunching instability. It occurs if the interaction of the bunch with its own radiation leads to the formation of sub-structures on the longitudinal bunch profile. These sub-structures can grow rapidly -- leading to a sawtooth-like behaviour of the bunch. At KARA, we use a fast-gated intensified camera for energy spread studies, Schottky diodes for coherent synchrotron radiation studies as well as electro-optical spectral decoding for longitudinal bunch profile measurements. For a synchronization, a hardware synchronization scheme is used which compensates for eventual hardware delays. In this paper, the different experimental setups and their synchronization are discussed and first results of synchronous measurements are presented.

Short-term acclimation response of individual cells of Thalassiosira weissflogii was monitored by Synchrotron FTIR imaging over the span of 75 minutes. The cells, collected from batch cultures, were maintained in a constant flow of medium, at an irradiance of 120 μmol m−2 s−1 and at 20 °C. Multiple internal reflections due to the micro fluidic channel were modeled, and showed that fringes are additive sinusoids to the pure absorption of the other components of the system. Preprocessing of the hyperspectral cube (x, y, Abs(λ)) included removing spectral fringe using an EMSC approach. Principal component analysis of the time series of hyperspectral cubes showed macromolecular pool variations (carbohydrates, lipids and DNA/RNA) of less than 2% after fringe correction.

A key bottleneck for investigations of ultrafast processes is a detection scheme to record all individual spectra with high repetition rates to avoid averaging and to improve the signal-to-noise ratio. Here, we present spectral measurements of fs laser sources used for electro-optical detection with a KIT-developed linear sensor array and DAQ system adapted to light sources with MHz repetition rates. This system can be equipped with different sensor types covering a broad wavelength range. It can therefore be used for various applications and scientific questions. The presented exemplary applications range from accelerator-based diagnostics to table-top laser experiments.

At the KArlsruhe Research Accelerator (KARA), we use electro-optical sampling to measures profiles of compressed electron bunches during the microbunching instability. The observation of the complex dynamics of this instability is of special interest because it leads to intense THz radiation bursts. As the revolution frequency of the storage ring is 2.72 MHz, high detection rates are required to record the bunch profiles for every revolution with single-shot measurements. To achieve fast detection rates, we implemented a KIT-developed ultra-fast line array and recorded the electron bunch charge density for every revolution for 3.6 s with a data throughput of 1.4 GBytes/s.

New methods are needed to rapidly identify malaria parasites in blood smears. The coupling of a Focal Plane Array (FPA) infrared microscope system to a synchrotron light source at IRENI enables rapid molecular imaging at high spatial resolution. The technique, in combination with hyper-spectral processing, enables imaging and diagnosis of early stage malaria parasites at the single cell level in a blood smear. The method relies on the detection of distinct lipid signatures associated with the different stages of the malaria parasite and utilises resonant Mie extended multiplicative scatter correction to pre-process the spectra followed by full bandwidth image deconvolution to resolve the single cells. This work demonstrates the potential of focal plane technology to diagnose single cells in a blood smear. Brighter laboratory based infrared sources, optical refinements and higher sensitive detectors will soon see the emergence of focal plane array imaging in the clinical environment.

We report Fourier transform infrared spectro-microtomography, a nondestructive three-dimensional imaging approach that reveals the distribution of distinctive chemical compositions throughout an intact biological or materials sample. The method combines mid-infrared absorption contrast with computed tomographic data acquisition and reconstruction to enhance chemical and morphological localization by determining a complete infrared spectrum for every voxel (millions of spectra determined per sample).

Using arbitrary periodic pulse patterns we show the enhancement of specific frequencies in a frequency comb. The envelope of a regular frequency comb originates from equally spaced, identical pulses and mimics the single pulse spectrum. We investigated spectra originating from the periodic emission of pulse trains with gaps and individual pulse heights, which are commonly observed, for example, at high-repetition-rate free electron lasers, high power lasers, and synchrotrons. The ANKA synchrotron light source was filled with defined patterns of short electron bunches generating coherent synchrotron radiation in the terahertz range. We resolved the intensities of the frequency comb around 0.258 THz using the heterodyne mixing spectroscopy with a resolution of down to 1 Hz and provide a comprehensive theoretical description. Adjusting the electron's revolution frequency, a gapless spectrum can be recorded, improving the resolution by up to 7 and 5 orders of magnitude compared to FTIR and recent heterodyne measurements, respectively. The results imply avenues to optimize and increase the signal-to-noise ratio of specific frequencies in the emitted synchrotron radiation spectrum to enable novel ultrahigh resolution spectroscopy and metrology applications from the terahertz to the x-ray region.

The 20th IEEE-NPSS Real Time Conference, RT2016, Padua, Italy, 5 Jun 2016 - 10 Jun 2016 ; 1-2(2016)., We developed an ultra-fast linear detector with a frame rate of up to 10 6 frames per second (fps) in order to improve the acquisition rate and the resolution of Electro-Optical Spectral Decoding (EOSD) setups currently installed at several light sources (ANKA, European XFEL, TELBE).The system consists of a detector board, an FPGA readout board and a high-throughput data link. The detector board mounts a single linear array sensor with 256 pixels and a 50 μ m pitch. InGaAs or Si sensors are used to detect near-infrared (NIR) or visible light. The GOTTHARD chip, developed at PSI, is used as charge-sensitive preamplifier. Four analog outputs, each one connected to 32 pixels through an analog multiplexer, operate at a maximum switching frequency of 33 MHz, resulting in a maximum frame rate of 10 6 fps.The analog outputs are digitized by a multi-channel ADC with 14 bits resolution. The data acquisition, the operation of the detector board and its synchronization with synchrotron machines are handled by the FPGA. The readout architecture is based on a PCI-Express high-throughput data link which allows continuous data acquisition at 450 MB/s when operating at a frame rate of 1 Mfps. The system is completed with a real-time data analysis module based on Graphics Processing Units (GPU) systems. We present the experimental measurements taken with a NIR laser operating at a repetition rate of 900 kHz, as well as the current performance in terms of frame rate and noise figures.

A new mid-infrared beamline (IRENI) extracting 320 hor. × 25 vert. mrads2 of radiation providing wide-field illumination for a Bruker Hyperion 3000 IR microscope equipped with an infrared focal plane array (FPA) detector has recently been commissioned at the Synchrotron Radiation Center (SRC) in Stoughton, WI. The swath of radiation from the SRC is extracted as 12 beams and recombined into a 3 × 4 bundle of beams that is refocused onto the sample plane of the infrared microscope illuminating a 40 μm × 60 μm sample area. With this new facility chemical images with high diffraction-limited resolution, for wavelengths across the mid-IR concurrently, can be obtained in minutes. Applications of biomedical, biological and cultural heritage interest show the broad applicability of this new facility. The examples presented here include images of fixed single cells, time dependent in vivo measurements of a single cell and layered organic/inorganic materials.

The critical questions into the cause of neural degeneration, in Alzheimer disease and other neurodegenerative disorders, are closely related to the question of why certain neurons survive. Answers require detailed understanding of biochemical changes in single cells. Fourier transform infrared microspectroscopy is an excellent tool for biomolecular imaging in situ, but resolution is limited. The mid-infrared beamline IRENI (InfraRed ENvironmental Imaging) at the Synchrotron Radiation Center, University of Wisconsin-Madison, enables label-free subcellular imaging and biochemical analysis of neurons with an increase of two orders of magnitude in pixel spacing over current systems. With IRENI's capabilities, it is now possible to study changes in individual neurons in situ, and to characterize their surroundings, using only the biochemical signatures of naturally-occurring components in unstained, unfixed tissue. We present examples of analyses of brain from two transgenic mouse models of Alzheimer disease (TgCRND8 and 3xTg) that exhibit different features of pathogenesis. Data processing on spectral features for nuclei reveals individual hippocampal neurons, and neurons located in the proximity of amyloid plaque in TgCRND8 mouse. Elevated lipids are detected surrounding and, for the first time, within the dense core of amyloid plaques, offering support for inflammatory and aggregation roles. Analysis of saturated and unsaturated fatty acid ester content in retina allows characterization of neuronal layers. IRENI images also reveal spatially-resolved data with unprecedented clarity and distinct spectral variation, from sub-regions including photoreceptors, neuronal cell bodies and synapses in sections of mouse retina. Biochemical composition of retinal layers can be used to study changes related to disease processes and dietary modification.

Spectroscopy and spatially resolved chemical imaging of biological materials using an infrared microscope is greatly enhanced with confocal image plane masking to 5–6 μm with a third generation microspectrometer and illumination with a synchrotron radiation source compared to globar illuminated and array detection or singly masked system. Steps toward this instrumental achievement are illustrated with spectra and images of biological tissue sections, including single cells, brain, aorta, and grain specimens. A recent, customized synchrotron infrared microspectrometer installation enables focal plane array detection to achieve both rapid and high definition chemical imaging. Localization of the ester carbonyl population in single modified starch granules was used to provide direct comparison of the two advanced imaging capabilities.

Table-top Fourier transform infrared (FT-IR) imaging using focal plane array (FPA) multi-element detectors is an increasingly popular chemical microscopy technique because it can provide microspectroscopic images of large sample areas in short times at moderate spatial resolution. The novel IR beamline IRENI at the Synchrotron Radiation Center (Wisconsin, USA), the first dedicated multi-beam synchrotron-based FT-IR imaging system, offers, within minutes, high quality chemical images at the highest available spatial resolution (diffraction-limited at all mid-IR wavelengths) with a pixel size of 0.54×0.54 μm 2 for transmission measurements. Due to this very high spatial sampling, mathematical image enhancement algorithms such as deconvolution and total variation (TV) reconstruction can be implemented to improve image contrast and thus spatial resolution. This is demonstrated for US Air force (USAF) targets, micron-sized aluminum beads, and a single living algal cell.

We have developed a liquid/flow cell/chamber allowing infrared measurements of living biological specimens with high spatial resolution under a controlled aqueous environment. This flow chamber features sub-micrometer thick diamond windows exhibiting low spherical and chromatic aberrations. Diamond has excellent transmission properties and minimal dispersion over the entire mid-infrared and visible spectral ranges. In contrast to current commercially available infrared liquid chambers, the flow chamber has a slim profile, which accommodates high resolution/magnification microscope objectives with small working distances, down to 0.6 mm above the chamber and 6 mm below the flow chamber. We have coupled a pump to the flow chamber to provide medium exchange. As an example, we present microspectroscopic infrared maps and spectra of the freshwater green alga Micrasterias sp. in the new flow chamber and compare them to maps obtained with a conventional liquid chamber. Pulse-amplitude-modulated fluorescence measurements on Micrasterias sp. cells inside the new flow chamber have been evaluated to demonstrate the viability of the algal cells.

University of Wisconsin—Milwaukee is designing and installing a mid-infrared beamline, IRMSI-MED, that will extract 320(h)×25(v) mrad 2 from a bending magnet (BM) at the Synchrotron Radiation Center. The BM radiation, collected with 12 toroidal mirrors and collimated with paraboloidal mirrors, illuminates a spot of 60×40 μm 2 at the sample plane of a commercial IR microscope. The microscope is equipped with a multi-element detector (MED) that will provide the opportunity to obtain chemical images with diffraction-limited resolution of the illuminated area in under a minute.

We present a method to realize active optical tips for use in near-field optics that can operate at room temperature. A metal-coated optical tip is covered with a thin polymer layer stained with CdSe nanocrystals or nanorods at low density. The time analysis of the emission rate and emission spectra of the active tips reveal that a very small number of particles - possibly down to only one - can be made active at the tip apex. This opens the way to near-field optics with a single inorganic nanoparticle as a light source.

We use fluorescent nanospheres as scalar detectors for the electric-field intensity in order to probe the near-field of an optical tip used in aperture-type near-field scanning optical microscopy (NSOM). Surprisingly, the recorded fluorescence images show two intensity lobes if the sphere diameter is smaller that the aperture diameter, as expected only in the case of vector detectors like single molecules. We present a simple but realistic, analytical model for the electric field created by light emitted by a NSOM tip which is quantitative agreement with the experimental data.

We report experimental measurements of the stability region of a Paul trap with very strong damping where the trapping parameter q was up to 25 times greater than the maximum value with no damping. We measured the region of stable trapping of charged particles for background pressures from atmospheric down to 5×10-2 mbar. The large increase in the size of the stability region that we observed at the highest pressure is explained by numerical solution of the Mathieu differential equation including a damping term. We show that stability regions that are separate under conditions of low damping merge together at higher pressures.

A new compact versatile linear accelerator named FLUTE is currently being designed at the Karlsruhe Institute of Technology. This paper presents the status of this 42 MeV machine. It will be used to generate strong (several 100 MV/m) ultra-short (∼1 ps) THz pulses (up to ∼4–25 THz) for photon science experiments, as well as to conduct a variety of accelerator studies. The latter range from comparing different coherent THz radiation generation schemes to compressing electron bunches and studying the electron beam stability. The bunch charge will cover a wide range (∼100 pC–3 nC). Later we plan to also produce ultra-short x-ray pulses from the electron bunches, which, for example, could then be combined for THz pump–x-ray probe experiments.

The wings of some insects, such as cicadae, have been reported to possess a number of interesting and unusual qualities such as superhydrophobicity, anisotropic wetting and antibacterial properties. Here, the chemical composition of the wings of the Clanger cicada (Psaltoda claripennis) were characterized using infrared (IR) microspectroscopy. In addition, the data generated from two separate synchrotron IR facilities, the Australian Synchrotron Infrared Microspectroscopy beamline (AS-IRM) and the Synchrotron Radiation Center (SRC), University of Wisconsin-Madison, IRENI beamline, were analysed and compared. Characteristic peaks in the IR spectra of the wings were assigned primarily to aliphatic hydrocarbon and amide functionalities, which were considered to be an indication of the presence of waxy and proteinaceous components, respectively, in good agreement with the literature. Chemical distribution maps showed that, while the protein component was homogeneously distributed, a significant degree of heterogeneity was observed in the distribution of the waxy component, which may contribute to the self-cleaning and aerodynamic properties of the cicada wing. When comparing the data generated from the two beamlines, it was determined that the SRC IRENI beamline was capable of producing higher-spatial-resolution distribution images in a shorter time than was achievable at the AS-IRM beamline, but that spectral noise levels per pixel were considerably lower on the AS-IRM beamline, resulting in more favourable data where the detection of weak absorbances is required. The data generated by the two complementary synchrotron IR methods on the chemical composition of cicada wings will be immensely useful in understanding their unusual properties with a view to reproducing their characteristics in, for example, industry applications.

Fourier transform infrared (FTIR) microspectroscopy is a powerful technique for label-free chemical imaging that has supplied important chemical information about heterogeneous samples for many problems across a variety of disciplines. State-of-the-art synchrotron based infrared (IR) microspectrometers can yield high-resolution images, but are truly diffraction limited for only a small spectral range. Furthermore, a fundamental trade-off exists between the number of pixels, acquisition time and the signal-to-noise ratio, limiting the applicability of the technique. The recently commissioned infrared synchrotron beamline, infrared environmental imaging (IRENI), overcomes this trade off and delivers 4096-pixel diffraction limited IR images with high signal-to-noise ratio in under a minute. The spatial oversampling for all mid-IR wavelengths makes the IRENI data ideal for spatial image restoration techniques. Here, we measured and fitted wavelength-dependent point-spread-functions (PSFs) at IRENI for a 74× objective between the sample plane and detector. Noise-free wavelength-dependent theoretical PSFs are deconvoluted from images generated from narrow bandwidths (4 cm(-1)) over the entire mid-infrared range (4000-900 cm(-1)). The stack of restored images is used to reconstruct the spectra. Restored images of metallic test samples with features that are 2.5 μm and smaller are clearly improved in comparison to the raw data images for frequencies above 2000 cm(-1). Importantly, these spatial image restoration methods also work for samples with vibrational bands in the recorded mid-IR fingerprint region (900-1800 cm(-1)). Improved signal-to-noise spectra are reconstructed from the restored images as demonstrated for a mixture of spherical polystyrene beads in a polyurethane matrix. Finally, a freshly thawed retina tissue section is used to demonstrate the success of deconvolution achievable with a heterogeneous, irregularly shaped, biologically relevant sample with distinguishing spectroscopic features across the entire mid-IR spectral range.

As silicon-based electronics are reaching the nanosize limits of the semiconductor roadmap, carbon-based nanoelectronics has become a rapidly growing field, with great interest in tuning the properties of carbon-based materials. Chemical functionalization is a proposed route, but syntheses of graphene oxide (G-O) produce disordered, nonstoichiometric materials with poor electronic properties. We report synthesis of an ordered, stoichiometric, solid-state carbon oxide that has never been observed in nature and coexists with graphene. Formation of this material, graphene monoxide (GMO), is achieved by annealing multilayered G-O. Our results indicate that the resulting thermally reduced G-O (TRG-O) consists of a two-dimensional nanocrystalline phase segregation: unoxidized graphitic regions are separated from highly oxidized regions of GMO. GMO has a quasi-hexagonal unit cell, an unusually high 1:1 O:C ratio, and a calculated direct band gap of ∼0.9 eV.

Conventional Fourier-transform infrared (FTIR) microspectroscopic systems are limited by an inevitable trade-off between spatial resolution, acquisition time, signal-to-noise ratio (SNR) and sample coverage. We present an FTIR imaging approach that substantially extends current capabilities by combining multiple synchrotron beams with wide-field detection. This advance allows truly diffraction-limited high-resolution imaging over the entire mid-infrared spectrum with high chemical sensitivity and fast acquisition speed while maintaining high-quality SNR.

We describe the fabrication of a corral trap, a new tool for the two-dimensional trapping of nanoscale particles, and present a software algorithm for automated trapping of a single particle and sequential trapping of multiple particles.

The use of synchrotron Fourier Transform Infrared spectroscopy (S‐FTIR) has been shown to be a very promising tool for biomedical research. S‐FTIR spectroscopy allows for the fast acquisition of infrared (IR) spectra at a spatial resolution approaching the IR diffraction limit. The development of the Infrared Environmental Imaging (IRENI) beamline at the Synchrotron Radiation Center (SRC) at the University of Wisconsin‐Madison has allowed for diffraction limited imaging measurements of cells in human prostate and breast tissues. This has allowed for the identification of cell types within tissues that would otherwise not have been resolvable using conventional FTIR sources.

First results from IRENI, a new beamline at the Synchrotron Radiation Center, demonstrate that synchrotron chemical imaging, which combines the characteristics of bright, stable, broadband synchrotron source with a multi‐element detector, produces diffraction‐limited images at all wavelengths simultaneously. A single cell of Micrasterias maintained in a flow cell has been measured, and results show high quality spectra and images demonstrating diffraction limited, and therefore wavelength‐dependent, spatial resolution.

Extended abstract of a paper presented at Microscopy and Microanalysis 2010 in Portland, Oregon, USA, August 1 – August 5, 2010.

The formation mechanism of the thermodynamically unstable calcite phase, very high Mg calcite, in biological organisms such as sea urchin or corallina algae has been an enigma for a very long time. In contrast to conventional methods such as KBr pellet Fourier Transform infrared (FTIR) spectroscopy and X-ray diffraction (XRD), FTIR microspectroscopy (FTIRM) provides additional information about a local disorder such as an amorphous phase or the occlusion of Mg ions in the calcite lattice. In this work, we characterise for the first time systematically synthetic and biogenic Mg-containing calcium carbonate samples (especially sea urchin teeth--SUT) in detail by using two FTIRM instruments and compare these samples with KBr pellet FTIR measurements. Furthermore, we present spectra from geogenic calcite and dolomite minerals, recorded with both FTIRM systems, as well as KBr pellet FTIR spectra as references. We analyse the spectra by applying multi-peak curve fitting on the in-plane-bending (ν(4)) and out-of-plane (ν(2)) bands. Based on the obtained results we attribute the two singlet bands at ∼860-865 cm(-1) and ∼695-704 cm(-1) observed in the SUT FTIRM spectra to the existence of amorphous calcium carbonate (ACC), and report for the first time the existence of ACC at the mature end of SUT. In the other three studied biominerals, however, we did not find any ACC. Also, based on the FTIRM results, we observe that not only ν(4), but also ν(2) shifts to higher wavenumbers if more calcium ions are replaced by magnesium ions in the calcite lattices.

In this paper we present a rigorous and general theoretical model for the illumination point spread function of a confocal microscope that correctly reproduces the optical setup. The model uses vectorial theory and assumes that monochromatic light with a Gaussian intensity distribution (such as from a laser or a single-mode fiber) is focused by a microscope objective with high numerical aperture and passes through stratified media on its way to the sample. This covers the important practical case of illumination through up to three layers, which is the situation most commonly encountered in biological microscopy (immersion oil, glass coverslip, aqueous sample medium). It also accounts for objectives that are corrected for a certain coverslip thickness and refractive index but operated under non-design conditions. Furthermore, illumination with linearly, circularly, or elliptically polarized light is covered by introducing a Babinet-Soleil compensator into the beam path. The model leads to a set of analytical equations that are readily evaluated. Two-dimensional intensity distributions for particular cases of interest are presented and discussed.

International audience; Fluorescent nanobeads with a diameter of 20 nm were used to map the three-dimensional point spread function in the near-focus region of a confocal microscope at high spatial resolution. Fluorescence images were taken in 109 equidistant planes (50 nm apart) parallel to the focal plane; postacquisition stacking of these images allows the reconstruction of the point spread function in the axial plane. The experimental distribution is compared to theoretical calculations based on an integral representation for the light intensity in the focus region that takes into account stratified media, polarization, the Gaussian illumination profile, and the finite exit pinhole size.