845 research outputs found

    Quantitative Imaging With DNA-PAINT for Applications in Synaptic Neuroscience

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    Super-resolution (SR) microscopy techniques have been advancing the understanding of neuronal protein networks and interactions. Unraveling the arrangement of proteins with molecular resolution provided novel insights into neuron cytoskeleton structure and actin polymerization dynamics in synaptic spines. Recent improvements in quantitative SR imaging have been applied to synaptic protein clusters and with improved multiplexing technology, the interplay of multiple protein partners in synaptic active zones has been elucidated. While all SR techniques come with benefits and drawbacks, true molecular quantification is a major challenge with the most complex requirements for labeling reagents and careful experimental design. In this perspective, we provide an overview of quantitative SR multiplexing and discuss in greater detail the quantification and multiplexing capabilities of the SR technique DNA-PAINT. Using predictable binding kinetics of short oligonucleotides, DNA-PAINT provides two unique approaches to address multiplexed molecular quantification: qPAINT and Exchange-PAINT. With precise and accurate quantification and spectrally unlimited multiplexing, DNA-PAINT offers an attractive route to unravel complex protein interaction networks in neurons. Finally, while the SR community has been pushing technological advances from an imaging technique perspective, the development of universally available, small, efficient, and quantitative labels remains a major challenge in the field

    Calculations of energy levels and lifetimes of low-lying states of barium and radium

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    We use the configuration interaction method and many-body perturbation theory to perform accurate calculations of energy levels, transition amplitudes, and lifetimes of low-lying states of barium and radium. Calculations for radium are needed for the planning of measurements of parity and time invariance violating effects which are strongly enhanced in this atom. Calculations for barium are used to control the accuracy of the calculations.Comment: 8 page

    Dual Magnetic Separator for TRIμ\muP

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    The TRIμ\muP facility, under construction at KVI, requires the production and separation of short-lived and rare isotopes. Direct reactions, fragmentation and fusion-evaporation reactions in normal and inverse kinematics are foreseen to produce nuclides of interest with a variety of heavy-ion beams from the superconducting cyclotron AGOR. For this purpose, we have designed, constructed and commissioned a versatile magnetic separator that allows efficient injection into an ion catcher, i.e., gas-filled stopper/cooler or thermal ionizer, from which a low energy radioactive beam will be extracted. The separator performance was tested with the production and clean separation of 21^{21}Na ions, where a beam purity of 99.5% could be achieved. For fusion-evaporation products, some of the features of its operation as a gas-filled recoil separator were tested.Comment: accepted by Nucl.Instr. Meth., final versio

    Radium single-ion optical clock

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    We explore the potential of the electric quadrupole transitions 7s\,^2S_{1/2} - 6d\,^2D_{3/2}, 6d\,^2D_{5/2} in radium isotopes as single-ion optical frequency standards. The frequency shifts of the clock transitions due to external fields and the corresponding uncertainties are calculated. Several competitive A^ARa+^+ candidates with A=A= 223 - 229 are identified. In particular, we show that the transition 7s\,^2S_{1/2}\,(F=2,m_F=0) - 6d\,^2D_{3/2}\,(F=0,m_F=0) at 828 nm in 223^{223}Ra+^+, with no linear Zeeman and electric quadrupole shifts, stands out as a relatively simple case, which could be exploited as a compact, robust, and low-cost atomic clock operating at a fractional frequency uncertainty of 10−1710^{-17}. With more experimental effort, the 223,225,226^{223,225,226}Ra+^+ clocks could be pushed to a projected performance reaching the 10−1810^{-18} level.Comment: 20 pages, 1 figur

    Super-resolution imaging and estimation of protein copy numbers at single synapses with DNA-PAINT

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    In the brain, the strength of each individual synapse is defined by the complement of proteins present or the "local proteome." Activity-dependent changes in synaptic strength are the result of changes in this local proteome and posttranslational protein modifications. Although most synaptic proteins have been identified, we still know little about protein copy numbers in individual synapses and variations between synapses. We use DNA-point accumulation for imaging in nanoscale topography as a single-molecule super-resolution imaging technique to visualize and quantify protein copy numbers in single synapses. The imaging technique provides near-molecular spatial resolution, is unaffected by photobleaching, enables imaging of large field of views, and provides quantitative molecular information. We demonstrate these benefits by accessing copy numbers of surface AMPA-type receptors at single synapses of rat hippocampal neurons along dendritic segments

    Determination of transition frequencies in a single 138^{138}Ba+^{+} ion

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    Transition frequencies between low-lying energy levels in a single trapped 138^{138}Ba+^{+} ion have been measured with laser spectroscopy referenced to an optical frequency comb. By extracting the frequencies of one-photon and two-photon components of the line shape using an eight-level optical Bloch model, we achieved 0.1 MHz accuracy for the 5d 2^{2}D3/2_{3/2} - 6p 2^{2}P1/2_{1/2} and 6s 2^{2}S1/2_{1/2} - 5d 2^{2}D3/2_{3/2} transition frequencies, and 0.2 MHz for the 6s 2^{2}S1/2_{1/2} - 6p 2^{2}P1/2_{1/2} transition frequency.Comment: 5 pages, 7 figures, submitted to Phys. Rev.

    Development of a thermal ionizer as ion catcher

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    An effective ion catcher is an important part of a radioactive beam facility that is based on in-flight production. The catcher stops fast radioactive products and emits them as singly charged slow ions. Current ion catchers are based on stopping in He and H2_2 gas. However, with increasing intensity of the secondary beam the amount of ion-electron pairs created eventually prevents the electromagnetic extraction of the radioactive ions from the gas cell. In contrast, such limitations are not present in thermal ionizers used with the ISOL production technique. Therefore, at least for alkaline and alkaline earth elements, a thermal ionizer should then be preferred. An important use of the TRIμ\muP facility will be for precision measurements using atom traps. Atom trapping is particularly possible for alkaline and alkaline earth isotopes. The facility can produce up to 109^9 s−1^{-1} of various Na isotopes with the in-flight method. Therefore, we have built and tested a thermal ionizer. An overview of the operation, design, construction, and commissioning of the thermal ionizer for TRIμ\muP will be presented along with first results for 20^{20}Na and 21^{21}Na.Comment: 10 pages, 4 figures, XVth International Conference on Electromagnetic Isotope Separators and Techniques Related to their Applications (EMIS 2007

    Review

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    Innovation in genomics, transcriptomics, and proteomics research has created a plethora of state-of-the-art techniques such as nucleic acid sequencing and mass-spectrometry-based proteomics with paramount impact in the life sciences. While current approaches yield quantitative abundance analysis of biomolecules on an almost routine basis, coupling this high content to spatial information in a single cell and tissue context is challenging. Here, current implementations of spatial omics are discussed and recent developments in the field of DNA-barcoded fluorescence microscopy are reviewed. Light is shed on the potential of DNA-based imaging techniques to provide a comprehensive toolbox for spatial genomics and transcriptomics and discuss current challenges, which need to be overcome on the way to spatial proteomics using high-resolution fluorescence microscopy
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