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LITERATUREHIGHLIGHTSliterature highlightsGold Nanodot Arrays for NanofabricationA large array of sub-10-nm sin-gle-grain gold nanodots for usein nanotechnology is describedby Nicolas Clément and col-leagues at the Institut d'Elec-tronique Microélectronique etNanotechnologie, CNRS, Univer-sity of Lille, France [Small7(18):2607-2613, 2011].A uniform array of single-grain gold nanodots, as small as5-8 nm, was be formed on siliconusing e-beam lithography. Theas-fabricated nanodots wereamorphous, and thermal annealing converted themto pure Au single crystals covered with a thin SiO2layer. These findings were based on physical mea-surements by AFM, atomic-resolution STEM, andchemical techniques using energy dispersive X-rayspectroscopy. The authors demonstrated the forma-tion by e-beam lithography of sub-10-nm Au dotswith small dispersion and perfect alignment. Suchprecise formation of small dots enabled them to iden-(a) STEM image showing the bulk silicon (Si), five annealed dots (Au), carbon layer (C), and platinum layers. (c) Coloured STEM image of a single annealed nanodot (260°C, 2 h). Reproduced with permission, Copyright ©2011 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim. tify the critical size that determines whether a dot iscomposed of single or multiple crystal domains.Moreover, they showed that annealing at moderatetemperature can convert Au dots from amorphous tosingle-crystalline, and then they were covered with athin SiO2layer. After easy removal of the SiO2(diluteHF etching), these nanodots can be used as electrodesfor the characterization of organic self-assembledmonolayers (SAMs) with less than 200 molecules.A Stage for Micro-AxialFluorescence TomographyA miniature stage device to overcome resolutionanisotropy in fluorescence light microscopy isdescribed by Florian Staier and colleagues at theKirchhoff Institute for Physics, University of Heidel-berg, Germany [Rev. Sci. Instrum. 82:093701, 2011].To overcome the limitation of fluorescence micro-scopes in anisotropic optical resolution or point local-ization precision micro-axial tomography was usedwhich allowed object tilting on the microscope stageand led to an improvement in localization precisionand spatial resolution. A glass fiber was placed in theobject space of the microscope lens and its rotationcontrolled by a miniaturized stepping motor. By Testparticles were fixed onto the glass fiber, opticallylocalized with high precision, and automaticallyrotated to obtain views from different perspectiveangles from which distances of corresponding pairs ofobjects were determined. From these angle depen-dent distance values, the real 3D distance was calcu-lated with a precision in the ten nanometer range(corresponding here to an optical resolution of 10-30nm) using standard microscopical equipment. As aproof of concept, the spindle apparatus of a maturemouse oocyte was imaged during metaphase II mei-otic arrest under different perspectives.Enabling Two-ColourSTED of Live CellsA technique for the use of two fluorophores in stim-ulated emission depletion (STED) microscopy of livingcells is reported by Patrina Pellett and co-workers atthe Department of Cell Biology, Yale School of Medi-cine, CT [Biomedical Optics Express2(8):2364-2371,2011]. Current applications of STED microscopy havebeen limited to single colour imaging of living cellsand multicolour imaging in fixed cells. However, tostudy active processes, such as protein interactions, atwo-colour STED imaging technique is needed in liv-ing cells. This was achieved for the first time by theauthors: the key to their success was in overcomingthe challenges in labeling target proteins in livingcells with dyes optimal for two-colour STEDmicroscopy. By incorporating fusion proteins, theresearchers were able to improve the targetingbetween the protein and the dye, effectively bridgingthe gap. This allowed the researchers to achieve reso-lutions of 78 nm and 82 nm for 22 sequential two-colour scans of epidermal growth factor and its recep-tor in living cells. 3D Point Spread Function in Cs-Corrected STEMThe three-dimensional point spread function of anaberration-corrected scanning transmission electronmicroscopy (STEM) has been simulated and experi-mentally tested by Andrew Lupini and Niels de Jongeat the Oak Ridge National Laboratory, TN [Microscopyand Microanalysis17:817-826, 2011].Aberration correction reduces the depth of field inSTEM and thus allows three-dimensional imaging bydepth sectioning. This imaging mode offers thepotential for sub-Ångstrom lateral resolution andnanometer-scale depth sensitivity. For biological sam-ples, which may be many ?m across and where highlateral resolution may not always be needed, opti-mizing the depth resolution even at the expense oflateral resolution may be desired, aiming to imagethrough thick specimens. Although there has beenextensive work examining and optimizing the probeformation in two dimensions, there is less knownabout the probe shape along the optical axis. The authors examined the probe shape in 3D in anattempt to better understand the depth resolution inImage Analysis of Deep Etched Actin FilamentsA fractal dimension analysis and mathematical mor-phology of structural changes in actin filamentsimaged by electron microscopy is reported by Yoshi-taka Kimori et al at the National Institutes of NaturalSciences, in Tokyo [Journal of Structural Biology176(1):1-8, Oct 2011].The authors examined structural changes of actinfilaments interacting with myosin visualized by quickfreeze deep-etch replica EM by using a new methodof image processing and analysis based on mathe-matical morphology. To quantify the degree of struc-tural changes, two characteristic patterns wereextracted from the EM images: the winding patternof the filament shape (WP) reflecting flexibility of theAtom Probe Imaging ofGaAsSb Quantum Rings A. Beltrán and colleagues at the Department of Mate-rials Sciences, University of Cadiz, Spain, report thatthree dimensional atom tomography resolves thequantum ring morphology of self-assembled GaSbburied nanostructures [Ultramicroscopy111(8):1073-1076, July 2011].Unambiguous evidence of ring-shaped self-assem-bled GaSb nanostructures grown by molecular beamepitaxy is presented on the basis of atom-probetomography reconstructions and darkfield transmis-sion electron microscopy imaging. From atom-probetomography compositional distribution has beenobtained. The GaAs capping process causes a strongsegregation of Sb out of the center of GaSb quantumdots, leading to the self-assembled GaAsxSb1-x quan-tum rings of 20-30 nm in diameter with x~0.33.filament, and the surface pattern of the filament (SP)reflecting intramolecular domain mobility of the actinmonomers in the filament. EM images were processedby morphological filtering followed by box-countingto calculate the fractal dimensions for WP (DWP) andSP (DSP). The result indicated that DWP was largerthan DSP irrespective of the state of the filament(myosin-free or bound) and that both parameters formyosin-bound filaments were significantly largerthan those for myosin-free filaments. This work is thefirst quantitative insight into how conformational dis-order of actin monomers is correlated with themyosin-induced increase in flexibility of actin fila-ments along their length.this mode. They present examples of how aberrationschange the probe shape in three dimensions, and itwas found that off-axial aberrations may need to beconsidered for focal series of large areas. It was shownthat oversized or annular apertures theoreticallyimprove the vertical resolution for three-dimensionalimaging of nanoparticles. When imaging nanoparti-cles of several nanometers in size, regular scanningtransmission electron microscopy can thereby be opti-mized such that the vertical full-width at half-maxi-mum approaches that of the aberration-correctedSTEM with a standard aperture.MICROSCOPY AND ANALYSIS 26NOVEMBER 2011

Free Educational Webinar: Sample Preparation for Scanning Electron Microscopy (SEM) State of the art Critical Point Drying with the New fully automated Leica EM CPD300Speaker: Dr. Ruwin Pandithage, Leica Mikrosysteme GmbH, Vienna, AustriaOnline Seminar/Webinar on Sample Preparation for SEM Register at http://www.microscopy-analysis.com/leicawebinars and join on Tuesday 25th October 2011 at 17:00 h (CET), 16:00 h (UK) Watch this webinar if you are responsible for . EM sample preparation of delicate biological specimens such as pollen, tissue, plants or insects. EM sample preparation of industrial samples for MEMS (Micro Electro Mechanical Systems) applications Specimens that can be damaged due to surface tension when changing from the liquid to gaseous state, need special treatment during sample preparation. Critical point drying is an efficient method for drying such delicate samples for SEM applications because it preserves the surface structure. Before drying, many biological samples are additionally prepared through fixation and dehydration and then coated after drying with a metal such as gold, platinum or palladium to make their surfaces electrically conductive for the SEM analysis. If the surface structure is altered during the drying process the results of the following SEM application will provide incorrect results.In the past, critical point drying was a time consuming process with low sample reproducibility due to manual operation.This webinar will show you how the New Leica EM CPD300 dries delicate biological or industrial samples in a fully automated and controlled process to preserve your samples for subsequent treatment and analysis.Register at http://www.microscopy-analysis.com/leicawebinarsThis webinar is available on demand under the above registration link.Critical Point Drying with Leica EM CPD300Free Educational Webinar: Sample Preparation for Scanning Electron Microscopy (SEM) State of the art Critical Point Drying with the New fully automated Leica EM CPD300Speaker: Dr. Ruwin Pandithage, Leica Mikrosysteme GmbH, Vienna, AustriaOnline Seminar/Webinar on Sample Preparation for SEM Register at http://www.microscopy-analysis.com/leicawebinars and join on Tuesday 25th October 2011 at 17:00 h (CET), 16:00 h (UK) Watch this webinar if you are responsible for . EM sample preparation of delicate biological specimens such as pollen, tissue, plants or insects. EM sample preparation of industrial samples for MEMS (Micro Electro Mechanical Systems) applications Specimens that can be damaged due to surface tension when changing from the liquid to gaseous state, need special treatment during sample preparation. Critical point drying is an efficient method for drying such delicate samples for SEM applications because it preserves the surface structure. Before drying, many biological samples are additionally prepared through fixation and dehydration and then coated after drying with a metal such as gold, platinum or palladium to make their surfaces electrically conductive for the SEM analysis. If the surface structure is altered during the drying process the results of the following SEM application will provide incorrect results.In the past, critical point drying was a time consuming process with low sample reproducibility due to manual operation.This webinar will show you how the New Leica EM CPD300 dries delicate biological or industrial samples in a fully automated and controlled process to preserve your samples for subsequent treatment and analysis.Register at http://www.microscopy-analysis.com/leicawebinarsThis webinar is available on demand under the above registration link.Critical Point Drying with Leica EM CPD300Online Seminar/Webinar on Sample Preparation for SEMRegister at http://www.microscopy-analysis.com/leicawebinarsCIRCLE NO. 25 OR ONLINE: www.microscopy-analysis.com