Thursday 01 September
Time Amphithéâtre Salle Bellecour 1,2,3 Salle Prestige Gratte Ciel Salle Gratte Ciel 1&2 Salon Tête d'Or Salle Gratte Ciel 3
08:45
08:45-09:45
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PL5
Plenary Lecture 5

Plenary Lecture 5

08:45 - 09:45 Plenary Lecture 5. Nadine PEYRIERAS (FRANCE)

10:15
10:15-12:30
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IM3-II
IM3: New Instrumentation
SLOT II

IM3: New Instrumentation
SLOT II

Chairmen: Emmanuel BEAUREPAIRE (Palaiseau, FRANCE), Christian COLLIEX (Orsay, FRANCE), Jörg ENDERLEIN (Göttingen, GERMANY), Andreas ENGEL (Delft, THE NETHERLANDS), Ernst H.K. STELZER (Professor) (Frankfurt am Main, GERMANY)
10:15 - 10:45 IM03-S39 Quantum coherent electron-light interactions studied by ultrafast transmission electron microscopy. Claus ROPERS (Göttingen, GERMANY)
Invited
10:45 - 11:00 #5203 - IM03-OP083 Coherence of a pulsed electron beam extracted from a semiconductor photocathode in transmission electron microscope.
Coherence of a pulsed electron beam extracted from a semiconductor photocathode in transmission electron microscope.

 Dynamic observations of nanoscale materials are important for investigations of the time evolution of optical couplings, distractions, and energy relaxations in a local site. To suppress electron-beam damage of biological specimens or organic material in transmission electron microscopes (TEMs), a pulsed electron beam is expected to be applied for the probe beam. Therefore, we have begun developing a spin-polarized pulse-TEM (SPTEM), which comprises a photocathode-type electron source (PES) and a low-voltage TEM [1-3]. Several beam parameters of the PES are greatly superior to those of conventional thermal electron beams. In addition, PES has the ability to generate a sub-picosecond pulse-beam [4].

 In continuous beam emissions, we have previously demonstrated that the SPTEM can provide both TEM images and diffraction patterns [2]. The TEM images were obtained at a spatial resolution of 1 nm with a 30-kV acceleration voltage. The apparatus has an electron beam energy width below 114-meV in the TEM without any monochrometors [6]. The energy width indicates that the temporal coherence is approximately 34 fs at 30-eV beam energy. The brightness is measured by taking a spot size and a convergent angle on an image plane. The measured brightness is approximately 4 × 107 A cm−2 sr−1 at 30-keV beam energy with a polarization of 82% and a drive-laser power of 800 kW/cm2 on the photocathode [6]. The brightness for 200-kV beam energy is estimated to be 3 × 108 A cm−2 sr−1, which is converted using a Lorentz factor. The order of the brightness is sufficient for an interference experiment. Figure 2 demonstrates interference fringes of a spin-polarized electron beam using a newly installed biprism. The resulting electron beam exhibits a long coherence length owing to its low initial emittance of 2.6 nm rad, which can generate interference fringes representative of a first-order correlation using an electron biprism. These results indicate that the SPTEM can provide enough coherence in both the lateral and longitudinal directions even if the semiconductor photocathode is used for an electron emitter.

 Pulse beam emission in the SPTEM was also performed using a combination of the semiconductor photocathode and an ultra-short pulse laser, which can realize a time-resolved measurement with the stroboscopic technique or the single-shot technique. The photocathode has high quantum efficiency on the order of 10−3 compared with other metal-type photocathodes, which can realized not only a continuous emission but also a pico-second pulse emission. The picosecond pulse duration was realized using a newly developed ultra-short pulse laser system, which comprises a mode-lock Ti-Sapphire laser, a compensator for group velocity dispersion, and a pulse-duration converter. Figure 2 shows a typical beam current measured using a Faraday-cup type current monitor. The repetition rate of the pulse beam is synchronized with a drive laser system. Time-resolved TEM imaging and pulsed interference fringes were also successfully conducted using a stroboscopic acquisition technique [7]. Figures 3a and b show the interference fringes using a continuous electron wave and a 20-ps pulsed electron wave under the same condition of the electron optics, respectively. In the continuous–mode, a 1-mA source current was used for the interference experiment. In contrast, the pulsed beam had a high charge of 150 fC/pulse with a repetition rate of 80 MHz, which is comparable with a 12-mA average current. Consequently, despite its high current density, the pulsed electron beam emitted from the photocathode has sufficient coherence to realize a time-resolved holography that can analyze phase information in a temporal space.

 The authors thank Drs. H. Shinada, M. Koguchi, and M. Tomita of the Hitachi Central Research Laboratory for fruitful discussions and encouragement. This research was supported by MEXT KAKENHI Grant Numbers 25706031 and 15K13404.

 

[1] M. Kuwahara et al., Appl. Phys. Lett. 101 (2012) 03310.

[2] M. Kuwahara et al., AMTC Letters 3 (2012) 180.

[3] M. Kuwahara et al., J. Phys.:Conf. Ser. 298 (2011) 012016.

[4] Y. Honda, et al., Jpn. J. Appl. Phys. 52, 086401-086407(2013).

[5] X.G. Jin et al., Appl. Phys. Express 1 (2008) 045002.

[6] M. Kuwahara et al., Appl. Phys. Lett. 105, 193101 (2014).

[7] M. Kuwahara et al., Microscopy 62, 607-614 (2013).

Makoto KUWAHARA (Nagoya, JAPAN), Kouta AOKI, Hiroshi SUZUKI, Hidefumi ASANO, Toru UJIHARA, Koh SAITOH, Nobuo TANAKA
11:00 - 11:15 #4759 - IM03-OP082 Design and realization of an ultrafast cold field emission source operating under high voltage.
Design and realization of an ultrafast cold field emission source operating under high voltage.

Investigation of nanostructures physics requires atomic spatial resolution, meV spectral resolution and femto to nanosecond time-resolution. Accessing all these informations simultaneously would be a breakthrough in nanophysics.

Ultrafast Transmission Electron Microscopes (UTEM) combining subpicosecond temporal resolution and nanometer spatial resolution have recently emerged as unique tools for investigations at both ultimate spatial and temporal resolutions [1]. However, the performances of state-of-the-art UTEM are, in practice, brightness limited by their ultrafast electron source. These sources are commonly based either on photocathode excited by an ultrafast laser beam [2] or, very recently, on Schottky type assembly [3].

The FemtoTEM project aims at developing an alternative Ultrafast Transmission Electron Microscope based on a high brightness laser-driven cold field emission electron source working under 200kV acceleration voltage. The latter consists of a metallic nanotip in tungsten illuminated by femtosecond laser pulses [4]. This development has been achieved by bringing together a commercial femtosecond laser source and a customized 200kV cold-field emission Transmission Electron Microscope Hitachi HF2000.

The main difficulty was to develop the ultrafast cold field emission source modifying the old HF2000 design, well known in CEMES [5]. Indeed, to perform a proper laser assisted field emission, the femtosecond laser beam need to be injected in the ultra high vacuum and high voltage area of the gun, where the tip is located, then focused and aligned in three dimensions on the fine apex of the FE W tip. The new design has been thought to allow such complex operation while keeping the possibility of cold field emitting electrons in continuous operation, as for the standard source. This new design has been finally produced and patented [6].

To build this new source, deep modifications, compared to the original Hitachi design, have been implemented, from the high voltage gun housing and cable to the inner structure of the gun assembly. With the help of finite element modeling, and ray tracing software, the influence of the new design on the electric field, and electrons trajectories, brightness, … has been investigated and compared to experimental results (see Figure). Last results will be also presented highlighting the potentiality of this new source for ultrafast electron holography application, for which a good brightness is mandatory. 

[1] Zewail, A. H., Science, 2010, 328, 187-193

[2] Zewail, A. H., USPTO n°US7,154,091 of December 26. 2006

[3] Bormann, R. et al, Journal of Applied Physics, 2015, 118, 173105

[4] Hommelhoff, P. et al, Phys. Rev. Lett., 2006, 96, 077401

[5] Houdellier, F. et al, Carbon 2012, 50 (5), 2037–2044

[6] Arbouet A. & Houdellier F., USPTO No.US9,263,229 B2 of February 16.2016

 

 Acknowledgments

This work was funded through the support of the « Institut National de Physique du CNRS »- INP-CNRS and the ANR FemtoTEM n°ANR-14-CE26-0013-01. The authors acknowledge the European Union under the Seventh Framework Programme under a contract for an Integrated Infrastructure Initiative Reference 312483-ESTEEM2.

Florent HOUDELLIER (TOULOUSE), Giuseppe Mario CARUSO, Pierre ABEILHOU, Arnaud ARBOUET
11:15 - 11:30 #5965 - IM03-OP089 Spatio-Temporal Probing of Lattice Dynamics in Graphite by Ultrafast TEM.
Spatio-Temporal Probing of Lattice Dynamics in Graphite by Ultrafast TEM.

Over the last decades, electron microscopy was tremendously successful in unravelling material structures and compositions, resolved on the atomic scale, but only with limited temporal resolution. Optical pump-probe techniques are now applied routinely for the study of ultrafast dynamics. Nevertheless, we still lack tools for accessing nanoscale dynamics on a femtosecond timescale.

Such a capability can be provided by ultrafast transmission electron microscopy (UTEM), which employs a pulsed electron beam with sub-picosecond pulse duration to stroboscopically probe ultrafast laser-driven dynamics with the imaging and diffraction capabilities of electron microscopy [1,2]. So far, the potential of this approach is limited by the availability of a high brightness laser-driven electron source within a transmission electron microscope.

Here, we apply UTEM for the study of ultrafast local lattice dynamics in single crystalline graphite, enabled by the generation of highly coherent electron bunches from a point-like photoelectron source [3].

The Göttingen UTEM instrument is based on the custom modification of a JEOL 2100F Schottky field emission TEM, allowing for optical sample excitation and the generation of optically triggered ultrashort electron pulses (Fig. 1a) [4]. The laser-triggered nanoscopic electron source [5-7] employs localized single-photon photoemission from the front facet of a tip-shaped ZrO/W(100) emitter (Fig. 1b). Highly coherent ultrashort electron pulses with a normalized emittance of 3 nm∙mrad are generated, enabling ultrafast electron imaging with phase-contrast and time-resolved local probing (Fig. 2). Specifically, at the sample position, we obtain electron focal spot sizes down to 1 nm with a temporal pulse width of 300 fs (full-width-at-half-maximum) and a spectral bandwidth of 0.6 eV (cf. Fig. 1c-e) [3].

We demonstrate ultrafast nanoscale diffractive probing, by studying the local light-induced structural dynamics close to the edge of a single-crystalline graphite thin film (Fig. 2a) [8]. Local convergent beam electron diffraction (CBED) patterns from nanoscale sample areas are recorded using tightly focused electron pulses (diameter of about 10 nm). The complex local distortion of the crystal structure is retrieved by utilizing the broad angular range of the incident electron beam (convergence angle of about 48 mrad) to probe several Bragg scattering conditions simultaneously in reciprocal space (cf. Fig. 2b,c).

For the case of graphite, we observe strongly pronounced lattice vibrations at the crystalline edge (Fig. 2d,e), corresponding to out-of-plane breathing modes, as well as in-plane shearing modes mapped with 10-nm spatial resolution. Considering the time-dependent relative line shifts, the individual contributions of mechanical deformation modes are disentangled. Furthermore, raster-scanning the electron focal spot across the sample allows for a comprehensive spatio-temporal reconstruction of the involved dynamics.

In conclusion, we have developed a novel UTEM instrument, relying on highly coherent electron pulses generated from a nanoscale photoemitter. Additionally, we presented first results on its capability for the investigation of ultrafast nanoscale dynamics in graphite.

References

[1] D. J. Flannigan, A. H. Zewail, Acc. Chem. Res. 45(10), 1828 (2012). [2] A. Yurtsever, A.H. Zewail, PNAS 108(8), 3152 (2011). [3] A. Feist et al., in preparation. [4] A. Feist et al., Nature 521, 200 (2015). [5] C. Ropers et al., Phys. Rev. Lett. 98, 043907 (2007). [6] M. Gulde et al., Science 345, 200 (2014). [7] R. Bormann et al., J. Appl. Phys. 118, 173105 (2015). [8] A. Feist et al., in preparation.

Armin FEIST (Göttingen, GERMANY), Nara RUBIANO DA SILVA, Wenxi LIANG, Claus ROPERS, Sascha SCHÄFER
11:30 - 11:45 #6771 - IM03-OP098 Chromatic corrected EFTEM investigation on spinodal decomposition of TiAlN at 80 kV with PICO.
Chromatic corrected EFTEM investigation on spinodal decomposition of TiAlN at 80 kV with PICO.

Chromatic aberration (Cc) in the modern transmission electron microscopy (TEM) plays an important role with the advancement the resolution up to atomic level1. Correction of Cc is especially important for high resolution energy -filtered TEM (EFTEM) with the requirement of large slit width and signal intensity to obtain an atomic resolved image, formatted from the inelastic scattered electrons in a limited time and stability constraint. As an advantage, large area and high resolution elemental map can be obtained in just tens of seconds, much faster than the traditional STEM-EELS counterpart. The short acquisition time is very crucial in the practical applications of both material science and biological science where the samples are very often sensitive to the electron beam illumination, the interesting compositions and structures are unknown, or they are the mixture of different elements and phases.

In this report, an annealed TiAlN sample is prepared for the EFTEM investigation with FEI-Titan PICO, focusing on the spinodal decomposition phenomenon. As a hard-coating material, TiAlN has exceptional mechanical and thermal properties attractive for tool-coating industry­­2. However at a higher temperature around 900 °C, the hardness is found to drop drastically owing to the formation of hexagonal AlN, which typically occurs after the spinodal decomposition. The spinodal decomposition contains the segregation of the composition while the crystal structure remains3. It is therefore typically difficult to prove the existence of this phenomenon without an elemental map with an atomic resolution, which is exactly a Cc corrected EFTEM image can provide.

In addition, different effects to the image resolution induced from factors considered in a Cc corrected EFTEM experiment will be discussed, including sample thickness, acceleration voltage, sample drifting, and atomic numbers of the sample. Compositional variation near the dislocation will be discussed as well. Preservation of elastic contrast4 has been observed and analyzed, which is crucial to map the inelastic image into the final elemental map.

 

Reference:

  1. K.W. Urban et al., Phys. Rev. Lett. 110, 185507 (2013).
  2. M. Hans et al., J. Appl. Phys. 116, 093515 (2014).
  3. I. Abrikosov et al., Materials 4, 1599-1618 (2011).
  4. A. Howie, Proc. R. Soc. Lond. A: Math. Phys Eng. Sci. 271, 268-287 (1963). 
Yen-Ting CHEN (AACHEN, GERMANY), Keke CHANG, Joachim MAYER, Jochen M. SCHNEIDER
11:45 - 12:00 #4572 - IM03-OP081 Status of the SALVE-microscope: Cc-correction for atomic-resolution TEM imaging at 20kV.
Status of the SALVE-microscope: Cc-correction for atomic-resolution TEM imaging at 20kV.

With the goal to enable atomic resolution TEM observations on beam sensitive materials the SALVE project had been initiated to develop a dedicated low-voltage TEM that is corrected for both, spherical and chromatic aberration [1,2].

The centerpiece of the SALVE III microscope is a new quadrupole-octupole Cc-Cs-corrector by CEOS that is based on the so-called Rose-Kuhn-Design [3,4]. The corrector is incorporated into a cubed FEI Titan Themis TEM and has been aligned for five accelerating voltages in the range from 20 to 80kV.

During design of the corrector special care had to be taken to prevent the resolution-limiting effect of thermal magnetic field noise (Johnson-Nyquist noise) that causes an image spread and limits the information transfer in Cc-corrected electron microscopes [5]. As seen in Figure 1(a), these measures have been very successful in that for all desired high-tensions the finally achieved resolutions exceed the 50mrad milestone. Figure 1(b) exemplarily demonstrates the achieved information limit on a purely amorphous 2nm tungsten sample at an accelerating voltage of 20kV. For the two shown perpendicular directions the Young's fringes significantly surpass the 50mrad aperture. In order to “use” the transferred information in a proper way, the phase plate, i.e. the aberration function, has to be well-controlled beyond the 50mrad-angle. This requires accurate control over axial aberrations up to including 5th order, and -for a considerable field of view- access to off-axial aberrations. The aberration measurement in Figure 1(c) demonstrates that all unround axial aberrations as well as the off-axial aberrations can be tuned sufficiently small. At the same time, the round aberrations can be adjusted for a suitable phase contrast transfer function (indicated in green color). Consequently, as shown in Figure 1(d) even at 20kV atomic resolution imaging becomes reality.

The chromatic aberration causes inelastically scattered electrons, i.e. electrons of lower energy, to be focused much stronger. This effect is compensated in the Cc-corrected instrument. Figure 1(e) compares the energy-dependent defocus effect of a Cc-uncorrected TEM (red line, Cc=1.45mm) and the SALVE microscope (measurements: black dots; 3rd order fit: dashed line) at 20kV. While the gradient of the Cc-uncorrected case is too steep to be distinguished from the axis of the ordinate in the magnified area, the SALVE instrument is capable to image a 20eV window with defocus changes of only 2nm. This will enable new imaging modes such as high-resolution EFTEM at very low voltages. [6]


References:

[1] U. Kaiser et al., Ultramicroscopy 111, Issue 8 (2011), 1239-1246.
[2] http://www.salve-project.de/
[3] H. Rose, Proc. 10th Eur. Congr. El. Micr. (Granada, Spain) (1992), 47.
[4] H. Rose, Patent Application DE 42 04 512 A 1 (1992).
[5] S. Uhlemann et al., Physical Review Letters 111(4) (2013), 046101.
[6] The authors acknowledge funding from the German Research Foundation (DFG) and the Ministry of Science, Research and the Arts (MWK) of the federal state Baden-Württemberg, Germany.

Martin LINCK (Heidelberg, GERMANY), Peter HARTEL, Stephan UHLEMANN, Frank KAHL, Heiko MÜLLER, Joachim ZACH, Johannes BISKUPEK, Marcel NIESTADT, Ute KAISER, Max HAIDER
12:00 - 12:15 #5987 - IM03-OP091 Aberration Corrected Analytical Scanning and Transmission Electron Microscope for High-Resolution Imaging and Analysis for Multi-User Facilities.
Aberration Corrected Analytical Scanning and Transmission Electron Microscope for High-Resolution Imaging and Analysis for Multi-User Facilities.

In recent years the revolution in aberration correction technology has made ultrahigh resolution imaging and analysis routinely accessible on transmission electron microscope (TEM) and scanning transmission electron microscope (STEM). We have developed a new analytical 200 kV cold field emission TEM equipped with a probe-forming aberration corrector, the model is Hitachi HF5000 (Figure 1). The microscope is fully covered in a metal enclosure to reduce the influence from environmental acoustic noise and temperature variation. Remote operation through Ethernet communication is possible as a result of a new design individual microprocessor circuit. Regarding the atomically resolved analytical capability, one of the key demands is to achieve high performance at a multi-user facility. To meet this demand, the Hitachi HF5000 is designed to be user friendly and extensive sample capability covers most requirement s from users in the fields of material science, materials fabrication, and device industry.

The HF5000 is capable of TEM imaging, STEM imaging with bright field (BF), annular dark field (DF) detectors, and secondary electron (SE) imaging. The probe-forming aberration corrector with automated correction of up to third order aberrations allows users to obtain aberration-free STEM illumination optics with minimized effort. Figure 2 gives an example after the aberration correction, the Ronchigram pattern of the amorphous specimen shows an approximately 32 mrad half angle flat region, corresponding to the optimal aperture condition for aberration-free STEM imaging. While TEM and STEM imaging probe the bulk structure of specimens, the SE imaging helps understanding the surface structure. It is important to note that the SE image can be acquired simultaneously with STEM image therefore both surface and bulk structures are revealed side-by-side at the same time, even at atomic resolution [1].  Such a triple imaging capability on one microscope column is very unique and critical in studying heterogeneous materials such as catalysts.

For modern analytical TEM, the most desirable features are high spatial resolution, high signal detection sensitivity, and sufficient specimen tilting angle range.  New pole-piece is therefore designed for HF5000 which enables sub-angstrom resolution DF-STEM imaging and a large specimen tilt angle of +/-35 degrees.  Figure 3 shows a high-angle annular DF (HAADF)-STEM image of a silicon thin film at the zone-axis direction, Si dumbbells with a 78 pm separation between adjacent Si columns are resolved. To make X-ray analysis highly sensitive, increasing solid angle of the Energy Dispersive X-ray spectroscopy (EDX) detector is essential. Two 100 mm2 silicon drift detectors (SDDs) are installed on the HF5000, the distance between detector and specimen (l) is carefully managed to maximize the solid angle. In order to increase peak to background ratio (P/B), the SDDs are positioned as high as possible relative to specimen surface.  The large take-off angle (q) reduces X-ray absorption and continuum X-rays which radiates with an angular intensity distribution. The improvement of resolving power is also related to the increased long term stability.  High voltage circuits are carefully designed, and stability for power supply of lens and deflection coils is enhanced. A stability of 0.6 ppm/min for circuit current is achieved for one of the beam deflection coils. In addition, the stability for the newly designed high tension circuit is less than 1 ppm which is critical to realize high energy resolution for analytical work using cold field emission gun and high spatial resolution for imaging.

[1] Zhu, Y., Inada, H., Nakamura, K. & Wall, J., Nature Mater. 8, 808-812 (2009)

Hiromi INADA (Ibaraki, JAPAN), Yoshifumi TANIGUCHI, Takafumi YOTSUJI, Keitaro WATANABE, Hirobumi MUTO, Wataru SHIMOYAMA, Hiroaki MATSUMOTO, Mitsuru KONNO
12:15 - 12:30 #6063 - IM03-OP093 Performances of aberration-corrected monochromatic low-voltage analytical electron microscope.
Performances of aberration-corrected monochromatic low-voltage analytical electron microscope.

To study the detailed electronic structures of carbon-related materials at an energy resolution better than 25 meV, we have developed a monochromatic low-voltage analytical electron microscope under a project “Triple-C phase-2”. The developed microscope is equipped with a double Wien-filter monochromator system [1] and delta-type aberration correctors [2].  It works at an accelerating voltage range from 15 kV to 60 kV.  The two Wien-filters for the monochromator, which is located just below the extraction anode of Schottky source, enables us to obtain an achromatic probe at an exit of the monochromator, since the second filter cancels out the energy dispersion generated by the first filter.  The energy spread of electrons (ΔE) is controllable by choosing the width of the slits, which are located between two filters, independently on the probe size at the specimen.  The delta-type aberration corrector consists of three dodeca-poles to correct geometrical aberrations up to the fifth-order including six-fold astigmatism.  In addition, so as to obtain high energy resolution spectra in electron energy-loss spectroscopy (EELS), the microscope is equipped with a high energy resolution spectrometer (Quantum-ERS from Gatan Inc.), which incorporates with the highly sensitive detection system at lower accelerating voltage and the highly stabilized power supplies for the prism and the lens system.

The ultimate energy resolution was acquired to be 14 meV at an accelerating voltage of 30 kV with an acquisition time of 2 ms, as shown in Fig. 1.  And at the longer acquisition time of 0.5 seconds, the energy resolution was measured to be 20 meV.  These results exceeded the original target of 25 meV.  Figures 2 (a) and (b) show raw ADF-STEM images of a single-layered graphene at 60 kV and 30 kV.  These images were obtained with a monochromatic electron probe, whose ΔE was 228 meV (using a 4 μm slit).  By using a monochromatic electron probe at 60 kV, C-C dumbbells of single-layered graphene were clearly resolved.  And a single-carbon atom on a graphene was successfully imaged at 30 kV.  These results indicate that the developed microscope enables us to analyze materials with high energy and spatial resolutions at lower accelerating voltage.

We tested a low-loss EELS map using a hexagonal boron nitride (h-BN) specimen with a monochromatic probe at 30 kV using a 0.1 μm slit. The experimental conditions are listed as probe size = 1 nm, probe current = 10 pA and acquisition time for each pixel = 0.3 seconds.   Figure 2 (a) shows the ADF-STEM image from the EELS mapping area. Fig. 2 (b) shows the extracted low-loss spectrum from the edge of the specimen indicated with the framed yellow square in Fig. 2 (a).  This spectrum, which was measured with an energy resolution of 22 meV, showed a sharp peak corresponding to an optical phonon at about 170 meV.  Figure 2 (c) shows the EELS map at the phonon excitation energy, whose intensity is normalized with the zero-loss peak intensity in each pixel. The phonon intensity was found to be strongly delocalized at the vacuum area beyond the edge of the specimen of more than 100 nm.

 

References

 

[1] M. Mukai, et al.: Ultramicroscopy 140 (2014) 37-43.

[2] H. Sawada, et al.: J. Electron Microsc. 58 (2009) 341-347.

[3] This work is supported by JST, Research Acceleration Program (2012–2016).

Masaki MUKAI (Akishima, JAPAN), Shigeyuki MORISHITA, Hidetaka SAWADA, Kazu SUENAGA

10:15-12:30
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IM8-II
IM8: Spectromicroscopies and analytical microscopy
SLOT II

IM8: Spectromicroscopies and analytical microscopy
SLOT II

Chairmen: Gerald KOTHLEITNER (Graz, AUSTRIA), Anders MEIBOM (Lausanne, SWITZERLAND), Bénédicte WAROT-FONROSE (Toulouse, FRANCE)
10:15 - 10:45 #8436 - IM08-S54 High resolution STEM imaging and analysis of 2D crystal heterostructure devices and nanoparticle catalysts.
High resolution STEM imaging and analysis of 2D crystal heterostructure devices and nanoparticle catalysts.

The new generation of aberration corrected scanning transmission electron microscope (STEM) instruments optimized for high spatial resolution energy dispersive x-ray (EDX) spectroscopy provide exciting opportunities for elemental analysis of nanoscale objects. Here I will discuss recent example applications from our studies of nanoparticle catalysts and 2D device heterostructures where these analytical capabilities have provided new insights to interpret the electronic and catalytic properties of such systems.

Elementally sensitive STEM EDX electron tomography provides a route to understanding full 3D morphology and chemistry with nanometre resolution. I will demonstrate results showing the effect of different elemental segregation on the catalytic performance of bimetallic nanoparticles [1]. I will also discuss the current limitations of this technique, including compensation of detector shadowing using a time varied acquisition scheme, and recent progress towards quantitative analysis [2,3].

I will also present work demonstrating that cross sectional STEM-EDX spectrum imaging can be used to reveal the internal atomic structure of van der Waals heterostructure devices produced by layering together multiple 2D crystals [4]. Recently we have studied light emitting diode devices, produced by mechanical exfoliation and subsequent stacking of 13 different 2D crystals, including 4 MoS2 monolayer quantum wells [5]. Using cross sectional STEM spectrum imaging we reveal that the crystal interfaces of such devices are atomically flat and provide detailed structural information to help to explain the electroluminescence results obtained. Other 2D crystal heterostructures will also be discussed including those incorporating air sensitive 2D crystals, such as black phosphorus, that require fabrication under an argon atmosphere to preserve the device performance [6].

Finally recent work where customised modification of an in situ STEM holder system [7] has allowed us to perform high spatial resolution STEM-EDX spectrum imaging during in-situ gas and liquid phase experiments and at elevated temperature [8].

 

[1] Slater et al, Nano Letters, 14, 1921-1926, (2014)

[2] Slater et al, Microscopy and Microanalysis, in press, (2016)

[3] Slater et al, Ultramicroscopy, 162, 61-73, (2016).

[4] Haigh et al, Nature Materials 11, 764-767, (2012); Georgiou et al Nature nanotechnology 8, 100-103 (2013)

[5] Withers et al, Nature Materials, 14, 301-306 (2015)

[6] Cao et al, Nano Letters, 15, 4914-4921 (2015)

[7] Zaluzec et al, Microscopy and Microanalysis 20 (2), 323 (2014)

[8] Lewis et al, Chemical Communications, 50, 10019-10022 (2014), Lewis et al Nanoscale, 6 (22), 13598-13605 (2014).

Sarah HAIGH (Manchester, UK), Thomas SLATER, Aidan ROONEY, Eric PRESTAT, Roman GORBACHEV, Freddie WITHERS, Konstantin NOVOSELOV, Geim ANDRE
Invited
10:45 - 11:15 #8364 - IM08-S54B Correlative investigations by HAADF-STEM and Atom Probe Tomography.
Correlative investigations by HAADF-STEM and Atom Probe Tomography.

The ultimate capabilities achieved by electron microscopies and their associated techniques inevitably raise the following question: is there room for conceiving new ways of investigating materials at the nano-scale? Indeed, most recent TEMs and STEMs easily achieve sub-Angström spatial resolution, while allowing elemental mapping at the same scale. Meanwhile, electron tomography has unambiguously demonstrated the possibility to image atomic positions and defects. In these instruments, some physical properties (e.g. optical, magnetic) are now accessible, again with increased resolution. However, as far as an ultimate machine would allow correlating physical properties with a “perfect” determination of atomic species and atomic positions in 3D, one must recognize that such a tool is not yet available. Aside from electron microscopes, Atom Probe Tomography (APT), which is intrinsically a 3D technique, has received increased attention owing to drastic developments during the last decade. This tool enables reconstructing volumes of matter by determining atom positions in 3D, which nature is determined by time of flight mass spectrometry. Thanks to the improvement of specimen preparation protocols, APT can be applied to much broader areas of materials science (semi-conductors, bio-materials, geo-materials, soft mater and even liquids).  Nowadays, intrinsic limitations of this tool reside in its limited detection efficiency (roughly 50% of atoms are detected) and in its anisotropic spatial resolution (though sub-Angström resolution is currently accessible along the direction of analysis, sub-nanometer resolution is achieved along transverse directions).

Strong advantages of APT rely in its possibility to detect all types of atoms, independently of their atomic number, in its excellent detection limit (few ppm in favorable cases but rarely more than 100 ppm), and in its intrinsic 3D nature. In order to collect a significant amount of information on a same nano-object, it is relevant to consider a correlative approach combining a TEM/STEM and APT. Motivations for such an approach are numerous. A non-exhaustive list would evoke: i) the possibility to associate structural defects (in TEM) with segregations (in APT); ii) associating the morphology of a particle (in electron tomography) with a 3D field of composition (in APT); iii) improving the quality of APT reconstructions by accessing additional information about the specimen morphology in TEM/STEM.

This presentation will begin with a rapid overview of the efforts made by the APT users community to promote this approach. Then, some illustrations will be given which relate the correlative investigation on alloys and quantum wells. The possibility to image APT specimens in STEM in high resolution mode (cf. Figure), while enabling atom counting will be discussed on the basis of HAADF-STEM image simulations. The advantages of the correlation of electron tomography and APT will be highlighted in the case of GaN/AlNGaN quantum wells.

Williams LEFEBVRE (ST ETIENNE ROUVRAY CEDEX), Florian MOYON, Antoine NORMAND, Nicolas ROLLAND, Ivan BLUM, Auriane ETIENNE, Celia CASTRO, Fabien CUVILLY, Lorenzo MANCINI, Isabelle MOUTON, Lorenzo RIGUTTI, François VURPILLOT
Invited
11:15 - 11:30 #5742 - IM08-OP141 Performance of the SALVE III corrector for EFTEM applications.
Performance of the SALVE III corrector for EFTEM applications.

The Sub-Angstroem-Low-Voltage-Electron-Microscope (SALVE) corrector was designed and built by the CEOS GmbH for the SALVE III project [1], a joined project of the group of Prof. Dr. Ute Kaiser at the University of Ulm (Germany), FEI company in Eindhoven (Netherlands) and CEOS GmbH (Germany). This Cc-Cs-corrector is a dedicated low-voltage corrector based on the so-called Rose-Kuhn-Design [2], operated in a cubed FEI Titan Themis TEM for acceleration voltages from 20kV to 80kV. For all these high tensions it can be aligned such that it provides uniform phase contrast transfer for all image features up to an aperture angle of θmax = 50mrad and at the same time for a considerable field of view.

 

To achieve such an excellent performance the corrector allows correcting all axial aberrations of fourth order and for certain unround axial aberrations of fifth order. Furthermore, C5 can be adjusted to its optimum positive value for bright atom contrast. All axial aberrations up to third order and all off-axial aberrations up to third order depending linearly on the distance from the axis can be adjusted. For all residual axial aberrations of fourth and fifth order, the lower order aberrations of same respective multiplicity can be adjusted for optimal compensation. This means that the integrated squared deviation from the ideal phase shift (±π/2 in case of C1, C3, C5 and zero for all other multiplicities) over the entire aperture is minimized for each azimuthal multiplicity separately [3]. The predicted performance of the corrector has already been demonstrated for the five acceleration voltages 20kV, 30kV, 40kV, 60kV and 80kV [4]. Figure 1 (a) shows that even within the largest field of view reasonable with the mounted Ceta 4k camera the wave aberration hardly changes up to a scattering angle of 50mrad.

 

Since the corrector will also be used together with a post column energy filter, we currently investigate both, theoretically and experimentally, how the optical performance of an EFTEM image is affected by the corrector. After correction of the linear chromatic aberration Cc, an energy window of at least 20eV can be transferred by the corrector with a negligible focus change even at a beam energy of 20keV, see figure 1 (b). However, there are many more potentially harmful types of chromatic aberrations to consider for an EFTEM image of a given finite width of the energy window:

 

   • Axial chromatic aberrations (depending on scattering angle and energy loss)

      can deteriorate the quality of the axial PCTF. For large EFTEM windows

      also the chromatic spherical aberration has to be taken into account.

   • Off-axial chromatic aberrations (depending on scattering angle, distance from the axis and energy loss)

      can effectively decrease the field of view, because they affect the quality of the transfer function

      in the outer image parts.

   • Chromatic distortions (depending on distance from the axis and energy loss)

      can deteriorate the information limit in the outer regions of an EFTEM image.

   • Residual dispersions of higher order in energy change could affect

      the information limit of all regions of the EFTEM image.

 

In this work we will analyze in detail how the residual higher order chromatic aberrations, the remaining chromatic distortions and the residual dispersions of the SALVE corrector affect the EFTEM performance. [5]

 

References:

 

[1] http://www.salve-project.de

[2] H. Rose, Proc. 10th Eur. Congr. El. Micr. (Granada, Spain) (1992), 47.

[3] M. Lentzen, Microsc. Microanal. 14 (2008)

[4] M. Linck: "Status of the SALVE-microscope", this conference.

[5] The authors acknowledge funding from the German Research Foundation (DFG) and the Ministry of Science, Research and the Arts (MWK) of the federal state Baden-Württemberg, Germany.

Frank KAHL (Heidelberg, GERMANY), Martin LINCK, Peter HARTEL, Heiko MUELLER, Stephan UHLEMANN, Max HAIDER, Joachim ZACH
11:30 - 11:45 #5760 - IM08-OP142 A new method for quantitative XEDS tomography of complex hetero-nanostructures.
A new method for quantitative XEDS tomography of complex hetero-nanostructures.

Over the last decades, electron tomography based on HAADF-STEM has evolved into a standard technique to investigate the morphology and inner structure of nanomaterials. The HAADF-STEM intensity depends on sample thickness but also scales with the atomic number Z and therefore, chemical compositions can be studied from these three-dimensional (3D) reconstructions.[1] Nevertheless, it is not straightforward to interpret the gray levels in a 3D HAADF-STEM reconstruction, when mixing of elements is expected or elements with atomic number Z close to each other are present.

In an increasing number of recent studies, X-ray Energy Dispersive Spectroscopy (XEDS) has been combined with tomography to understand complex nanostructure morphology and composition in 3D.[2] These studies rely on newly developed XEDS detectors such as the Super-X detection system, which consists of four individual detectors, symmetrically arranged around the TEM sample.[3] By using the Super-X detector, one is able to overcome problems that were previously related to extreme shadowing of the XEDS signal caused by the sample-detector configuration. Although this problem can be largely overcome, some shadowing effects remain, as illustrated in Figure 1. Since such shadowing effects vary for different tilt angles, the XEDS will also depend on the tilt angle and the projection principle for electron tomography is no longer fulfilled. Because of this problem and the low signal-to-noise ratio, typical of XEDS mapping, it remains challenging to obtain quantitative information by 3D XEDS and further progress is required.

Here, we propose an alternative approach to optimize the reconstruction of an XEDS tomography series by minimizing the impact of shadowing effects and improving the spatial resolution. The method is based on the synergistic combination of HAADF-STEM tomography and XEDS quantitative mapping.[4-5] HAADF-STEM yields a relatively high signal-to-noise ratio, enabling an accurate reconstruction of the morphology. XEDS, on the other hand, yields chemical information, but the limited amount of data that can be usually collected, hampers a good morphological reconstruction. As a proof of principle, we apply our methodology to a nanostructure containing a mix of Au and Ag atoms. It should be mentioned that the approach we propose here enables quantitative 3D chemical characterization of a broad variety of nanostructures.


 

[1]          P. Midgley, M. Weyland, Ultramicroscopy 2003, 96, 413.

[2]          B. Goris, L. Polavarapu, S. Bals, G. Van Tendeloo, L. M. Liz-Marzán, Nano letters 2014, 14, 3220.

[3]          P. Schlossmacher, D. Klenov, B. Freitag, H. Von Harrach, Microscopy today 2010, 18, 14.

[4]          D. Zanaga, T. Altantzis, J. Sanctorum, B. Freitag, S. Bals, Ultramicroscopy 2016, In press, doi:10.1016/j.ultramic.2016.03.002.

[5]          D. Zanaga, T. Altantzis, L. Polavarapu, L. M. Marzán, B. Freitag, S. Bals, Particle & Particle Systems Characterization, 2016, In press.

 

Acknowledgements

The authors acknowledge financial support from European Research Council (ERC Starting Grant # 335078-COLOURATOMS, ERC Advanced Grant # 291667 HierarSACol and ERC Advanced Grant 267867 - PLASMAQUO), the European Union under the FP7 (Integrated Infrastructure Initiative N. 262348 European Soft Matter Infrastructure, ESMI and N. 312483 ESTEEM2).

Daniele ZANAGA (Antwerpen, BELGIUM), Thomas ALTANTZIS, Lakshminarayana POLAVARAPU, Luis M. LIZ-MARZÁN, Bert FREITAG, Sara BALS
11:45 - 12:00 #5979 - IM08-OP147 X-ray emission generation constant from mono-layer graphene measured using STEM-EDS map detected with highly sensitive EDS system.
IM08-OP147 X-ray emission generation constant from mono-layer graphene measured using STEM-EDS map detected with highly sensitive EDS system.

X-ray generation constant of an atom by an electron irradiated is essential information for quantification of sample in X-ray fluorescence spectroscopy. The generated X-ray intensity (I) from a sample is described as I = 1/A ʃAʃt P(x, z)Ik(x) dzdx,  where A is electron beam irradiated area, t is sample thickness, P is electron density and Ik is X-ray intensity excited by an electron. To evaluate I, it is requested the accurate value of Ik. Then, to have accurate Ik, the comparison between Ik values from theoretical calculation (Ik theory) and experimental result (Ik exp) is important.  However, it is exacting to measure the Ik exp accurately using a multi-layer specimen because of two difficulties, especially for low energy X-rays. One is self absorption of emitted X-ray, which can be judged from the 3D sample shape. And the other is counting a number of atoms included in an analysis area. Namely, it is difficult to grasp the 3D sample shape. The shapes of single atomic chain or single layer specimen are simple and ideal for the aimed experiment in this paper. In this paper, we report the way and results of directly measured X-ray generation constant of carbon atoms, using a mono-layered graphene sample, which is easy to estimate 3D shape from electron microscope images.

For experiment, highly sensitive detector system is requested, since the X-ray signal from this sample is so thin and emit a little of X-ray. We used a multiple silicon drift detector (SDD) system for the X-ray measurement, since the SDD has become highly sensitive recently due to design flexibility of its shape and size to fit busy space around the sample of TEM. The microscope we used was an aberration corrected microscope (JEM-ARM300F).  For the new detection system with two SDD , a pair of objective lens pole pieces and a sample holder are re-designed to make a distance between sample and detectors as short as possible, and each detector size is enlarged to be 100 mm2, resulting in the total detection solid angle of 1.6 sr.

Figure 1(c) shows the highly magnified ADF image of mono-layer graphene from the square indicated by dashed line shown in Fig. 1(a). From the image we can recognize the area of mono-layer graphens by clear lattice image and hole of the sample. An X-ray elemental map shown in Fig. 1(b) was obtained simultaneously with ADF image shown in Fig. 1(a).  The difference of X-ray intensity between the hole and the mono-layer graphene is apparent in Fig. 1(b). This leads that the X-ray signal from mono layer with dual SDD detection system is well detectable under the dose, at which mono-layer graphene maintains its structure. The experimental conditions for this experiment were followings: accelerating voltage = 80 kV, probe current = 169 pA, number of pixels = 128 x 128 and total acquisition time = 300 sec.

The Ik exp of carbon is estimated to be 3.29×10-23 photons*cm2/electrons, using the experimental conditions and X-ray intensity from the mono-layer graphene area on Fig. 1(b). This is described as Ik exp ≈ 4πItotal/(Dcarbon*S*Delectron*W), where Itotal  is total X-ray counts (photons) from scan area of mono-layer graphene, Dcarbon is density of carbon atoms (atoms/cm2) in a analyzed area, S is scan area of graphene (cm2), W is a solid angle of X-ray detection and Delectron is density of electrons (electrons/cm2). Dcarbon was estimated to be 3.82×1015 atoms/cm2 from typical atomic density for a unit area. S is 1.38×10-13 cm2. The amount of X-ray counts (Itotal) was measured to be 155 photons accumulated over the area of mono-layer graphene. The density of electrons (Delectron) is simply calculated to be 7.22×1022 electrons/cm2 from experimental conditions. The theoretical generation constant (Ik theory) was calculated to be 1.85×10-23 photons*cm2, which was calculated as a product of ionization cross section (ionization atoms*cm2/electrons) by Bethe [2] and X-ray fluorescence yield (photons/ionization atoms) by Burhop [3], which is shown as Ik theory σkWk , where σk is ionization cross section and calculated to be 2.70×10-20 ionization atoms*cm2 for carbon, Wk is x-ray fluorescence yield and calculated to be 7.11×10-4 photons/ionization atoms for C Ka. The experimental value (Ik exp) do not differ from one by pure calculation (Ik theory) so much. The difference may be due to error in the pure calculation, since the calculation contains considerable approximate expressions. In conclusion, with the highly sensitive detection system and 2D sheet sample, we could measure a physical constant with considerable accuracy. The similar 2D samples such as BN and MoS2 sheets should be useful for measuring X-ray generation constants.

 

[Reference]

[1] NR Lugg et al,Ultramicroscopy 151(2015),p. 150.

[2] H Bethe, Ann. Physik  397(1930),p. 325.

[3] EHS Burhop, J. Phys. Radim 16(1955),p. 625. 

Yu JIMBO (3-1-2, Musashino, Akishima, Tokyo, JAPAN), Takeo SASAKI, Hidetaka SAWADA, Eiji OKUNISHI, Yukihito KONDO
12:00 - 12:15 #6368 - IM08-OP155 Correlative Nanoscale Luminescence and Elemental Mapping in InGaN/(Al)GaN Dot-in-a-wire Heterostructures.
Correlative Nanoscale Luminescence and Elemental Mapping in InGaN/(Al)GaN Dot-in-a-wire Heterostructures.

Ternary InGaN compounds show great promise for light-emitting diode (LED) applications because of bandgap energies (0.7 – 3.4 eV) that can be tailored to have emission wavelengths spanning the entire visible spectrum. Complex III-N device heterostructures have been incorporated into GaN nanowires (NWs) recently, but exhibit emission linewidths that are broader than expected for their corresponding planar counterparts, as measured with photoluminescence (PL) spectroscopy [1]. Nanoscale elemental mapping has provided evidence of alloy non-uniformity in NWs as a likely cause [2]. It is thus critical to understand how the structural and optical properties interplay within individual NW structures, using combined spectroscopic methods that can resolve different localized signals at the nanoscale with analytical scanning transmission electron microscopy (STEM).

 

Multiple InGaN/(Al)GaN quantum dot (QD) embedded nanowire heterostructures (NWHs), grown catalyst-free on Si(111) substrates by molecular beam epitaxy, were characterized by STEM. To investigate the inhomogeneous broadening observed in PL from an ensemble of NWs [1], nanometer-resolution STEM-cathodoluminescence (CL) spectral imaging on single NWs was performed at 150 K using a system as described in [3]. Individual NWs examined show diverse optical responses, but most NWs exhibit multiple sharp emission peaks (25 – 50 nm at FWHM) centered between 500 – 625 nm in the yellow-green wavelengths (Fig. 1i) from the active region (Fig. 1b–d), identified using the annular dark-field (ADF) signal collected concurrently. This is consistent with the PL, indicating that the broad emission originates from within single NWs. Subsequent aberration-corrected STEM-HAADF images on the same NWs were acquired to evaluate their structural properties, such as the size and morphology of the 10 QDs within the NWH (Fig. 1f). Additionally, electron energy-loss spectroscopy (EELS) spectrum imaging (SI), together with multiple linear least-squares fitting, was used to extract the In-distribution to quantify the In-composition projected through thickness (Fig. 1g,h) [2]. Apparent spatial-spectral correlation can be made between shifts in the CL emission wavelength to the relative In-content between successive QDs from the STEM-EELS (Fig. 1h, regions are color-coded to the corresponding emission wavelength based on the legend in Fig. 1i inset).

 

The luminescence intensity within NWs is related to the presence of a GaN or AlGaN shell surrounding the InGaN/GaN NWHs, formed due to sidewall incorporation during the growth of the subsequent GaN barrier and p-AlGaN electron-blocking layer (EBL), respectively. Both can enhance the in-plane confinement of carriers, hence reducing non-radiative surface recombination. Therefore, the utilization of larger bandgap AlGaN as barriers were also investigated for their expected enhancement in carrier confinement [4]. The InGaN/AlGaN NWHs exhibit a nested core-shell structure made up of an Al-rich AlGaN shell surrounding the InGaN QDs along axial and radial directions (Fig. 2g). STEM-CL spectral imaging shows a progressive red-shifting of the emission peaks along the growth direction (Fig. 2b–d,f,h). Spatial localization of individual spectral features suggests superior three-dimensional carrier confinement, which can be assigned to specific QDs as resolved in the bright-field (BF) image (Fig. 2a). Lastly, the observed spatial asymmetry in the luminescence intensity distribution, which is affected by charge carrier diffusion and drift in the presence of spontaneous and piezoelectric polarization fields in the InGaN/(Al)GaN NWHs, will also be addressed [5].

 

[1] H.P.T. Nguyen et al., Nano Lett., 12(3), 1317–1323 (2012)

[2] S.Y. Woo et al., Nanotechnology, 26(34), 344002 (2015); S.Y. Woo et al., Nano Lett., 15(10), 6413–6418 (2015)

[3] L. Zagonel et al., Nano Lett., 11(2), 568–573 (2011); L. Tizei et al., Appl. Phys. Lett., 105(14), 143106 (2014)

[4] H.P.T. Nguyen, M. Djavid, S.Y. Woo et al., Sci. Rep., 5, 7744 (2015)

[5] This work was supported by the Natural Sciences and Engineering Research Council of Canada (NSERC), with additional funding from the Michael Smith Foreign Study Supplement (MSFSS).

Steffi Y WOO (Hamilton, CANADA), Mathieu KOCIAK, Hieu P T NGUYEN, Zetian MI, Gianluigi A BOTTON
12:15 - 12:30 #6573 - IM08-OP159 How the Detector Geometry Influences EDXS Quantification.
How the Detector Geometry Influences EDXS Quantification.

Thanks to the recent advent of very large area sensors and multi-sensor systems X-ray spectrometry in analytical electron microscopes received a substantial boost. In particular, the combination of 2 or 4 detectors has proven to obtain such a good sensitivity that the detection of single atoms is feasible [1]. However, if it comes to quantitative work partial or full shadowing of one or more detectors by the specimen holder will introduce systematic errors of the obtained results [2].

In this paper we closely investigate the influence of the detector-microscope geometry on quantification using a ChemiSTEM system (Super-X) on a Titan microscope with a low background double tilt high visibility holder [3]. We introduce a procedure, which allows for the first time to determine experimentally the detector positions inside the microscope chamber in terms of elevation angle, azimuthal angle, and distance. We measured the amount of shadowing as a function of specimen tilt angle α and compared these measurements to simulations, where we varied the detector positions in order to optimize the fit to the experimental data. The sample was a specially designed Au-Pd thin film deposited on a silicon nitride membrane grid. The obtained positions were then verified by comparing the results to data obtained from different positions of the specimen holder.

To measure the amount of shadowing we used normalized intensities of the available X-ray lines. An optimized simulation for one of the Super-X detectors (Q1) may be seen in fig. 1 (Si-K data). By comparing measurements from different lines we noted a significant difference depending on the X-ray energies (compare Si-K and Au-L data in fig. 1). By further investigating this issue we found that the Beryllium specimen carrier inside the specimen holder may turn transparent for high enough X-ray energies (e.g. Au-L). In other words, the amount of shadowing not only depends on tilt angles and specimen positions but also on the energy of the X-ray line of interest (see fig. 2).

Once the detector positions are known the shadowing effects can be taken into account and their influence on quantitative results can be corrected. In this paper we characterize potential effects and systematic errors they introduce, which are relevant to conventional EDXS work as well as analytical tomography experiments. The procedures described will transform EDXS on a multi-detector system from qualitative analysis to full quantitative analysis.

 

Acknowledgements:

We greatly acknowledge the help of Dr. Bernd Oberdorfer, Austrian Foundry Research Institute (ÖGI), Leoben, Austria for performing the X-ray tomography experiments, which were funded by the Austrian Research Promotion Agency (FFG) (project OptimatStruct 839958). The authors also want to thank the Nanoinitiative Austria (Austrian Research Promotion Agency, FFG) for financial support as part of this work was done in the NILaustria cluster (project NILecho II 830269).

 

[1] T. C. Lovejoy, Q. M. Ramasse, M. Falke, A. Kaeppel, R. Terborg, R. Zan, N. Dellby, and O. L. Krivanek, “Single atom identification by energy dispersive x-ray spectroscopy,” Appl. Phys. Lett., vol. 100, no. 15, p. 154101, 2012.

[2] T. J. A. Slater, A. Janssen, P. H. C. Camargo, M. G. Burke, N. J. Zaluzec, and S. J. Haigh, “STEM-EDX tomography of bimetallic nanoparticles: A methodological investigation,” Ultramicroscopy, vol. 162, pp. 61–73, 2016.

[3] P. Schlossmacher, D. O. Klenov, B. Freitag, H. S. von Harrach, and A. Steinbach, “Nanoscale chemical compositional analysis with an innovative S/TEM-EDX system,” Microsc. Anal., vol. 24, no. 7, pp. 5–8, 2010.

Johanna KRAXNER, Margit SCHÄFER, Otto RÖSCHEL, Manuel PALLER, Georg HABERFEHLNER, Gerald KOTHLEITNER, Werner GROGGER (Graz, AUSTRIA)

10:15-12:30
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MS6-III
MS6 : Oxide-based, Magnetic and other Functional materials
SLOT III

MS6 : Oxide-based, Magnetic and other Functional materials
SLOT III

Chairmen: Etienne SNOECK (Toulouse, FRANCE), Maria VARELA (Madrid, SPAIN)
10:15 - 10:45 #8367 - MS06-S83 Split-illumination Electron Holography Applied to Electrostatic Potential Analyses of Oxide Heterojunctions with Polar Discontinuity.
Split-illumination Electron Holography Applied to Electrostatic Potential Analyses of Oxide Heterojunctions with Polar Discontinuity.

   Electron holography using interference of electron wave is one of quantitative microscopic techniques to visualize electromagnetic fields at nanometer scale. The long standing problem in this method was that the observable area was limited near the sample edge. To solve this problem, we developed split-illumination electron holography [1]. In this method, a coherent electron wave is separated into two coherent waves (object and reference waves) using biprism placed in the illumination system. The coherence degree of these electron waves do not change when they are separated. This makes it possible to achieve high precision holographic observation of an area far from the sample edge. As one of applications, electrostatic potential analysis of oxide heterojunctions with polar discontinuity will be presented.

   The discovery of the two-dimensional interface conduction in the LaAlO3/SrTiO3 (LAO/STO) heterojunction with polar discontinuity made a revival of the interest on the polar interface [2]. The emergence of high-mobility in LAO/STO is generally explained by the polar catastrophe scenario. Although the spontaneous electric polarization due to this charge redistribution was theoretically predicted, the existence of the mobile charges made it difficult to confirm the spontaneous polarization in LAO/STO junction. We explored oxide heterojunctions where the spontaneous polarization plays a dominant role in the charge screening and selected the LaFeO3 (LFO)/STO as target interface. Figure 1 shows two types of atomic sequences at the interfaces and reversal of electrostatic potential slopes in LFO. The results indicate that the originally non-polar LFO are converted into polar as a consequence of the polar catastrophe [3].

 

References

[1] T. Tanigaki, Y. Inada, S. Aizawa et al., Appl. Phys. Lett. 101, 043101 (2012).

[2] A. Ohtomo and H. Y. Hwang, Nature 427, 423 (2004).

[3] M. Nakamura, F. Kagawa, T. Tanigaki et al., Phys. Rev. Lett. 116, 156801 (2016).

 

Acknowledgements

This work was partly supported by the Japan Society for the Promotion of Science (JSPS) through its Funding Program for World-Leading Innovative R&D on Science and Technology (FIRST Program), Grant-in-Aid for Young Scientists (A) (15H05426) and that for Scientific Research (24226002) from the MEXT of Japan, and CREST from Japan Science and Technology Agency (JST).

Toshiaki TANIGAKI (Saitama, JAPAN), Masao NAKAMURA, Fumitaka KAGAWA, Hyun Soon PARK, Tsuyoshi MATSUDA, Daisuke SHINDO, Yoshinori TOKURA, Masashi KAWASAKI
Invited
10:45 - 11:00 #6045 - MS06-OP284 Complex magnetic distribution of diameter-modulated FeCoCu nanowires resolved by Electron Holography.
Complex magnetic distribution of diameter-modulated FeCoCu nanowires resolved by Electron Holography.

In the last years, diameter-modulated (D-M) ferromagnetic nanowires (NWs) have been intensively studied to evaluate their efficiency to control the motion of domain walls (DWs) along these one-dimensional nanostructures by the application of magnetic field or the injection of electrical current, which is essential for spintronic applications in the field of information storage, sensors and logical operations [1]. Preliminary studies in D-M NWs have been performed using theoretical and experimental procedures [2-4] in individual and isolated NWs and they have provided a first approach of the spin configuration in these systems, obtaining a non-trivial interpretation.

In this work, we have exploited the potential of electron holography technique (high spatial resolution, high sensitivity and quantitative capability in volume) for achieving a full picture of the magnetic distribution in cylindrical D-M FeCoCu NWs. These NWs were prepared by filling self-assembled cylindrical D-M nanochannels of anodic aluminum oxide templates. The D-M geometry of the polycrystalline NWs consist of alternating segments of small (100 nm) and large (144 nm) diameters, with segment lengths ranging between 1000 to 300 nm. At remanence, the high-shape anisotropy of the NWs induces a single-domain state where the spins are mainly oriented along the NW axis, with the possibility to create a small closure domain in large-diameter tips. The transition zones where the diameter is varied induce a complex demagnetizing field where the stray field follows a flux-closure configuration around the large-diameter segments and a magnetic coupling between them around small-diameter segments (See Fig 1). The complex configuration of the demagnetizing field can be understood if we treat the D-M transition zones as magnetic charges. The interpretation of the magnetic distribution by EH experiments was compared with micromagnetic simulation finding a very good agreement (see Fig 2). In addition, In-situ Lorentz microscopy experiments of the magnetization reversal process allowed evaluating the DW nucleation and propagate process by the application of magnetic fields. We found that a field-driven manipulation of DWs is not possible for the NWs in study. This “unsuccessful” result however helps us to take a step forward for the optimization of the geometry to reach the desired DW manipulation.

 

 

References

[1] Magnetic Nano- and Microwires edited by M. Vázquez. Woodhead Publising Series in Electronic and Optical Materials (2015).

[2] F. Tejo et al. Journal of Applied Physics 115 (2014) 17D136

[3] E. M. Palmero et al. Nanotechnology 26 (2015) 461001

[4] O. Iglesias-Freire et al. Nanotechnology 26 (2015) 395702

 

Acknowledgements

This work has been supported by ESTEEM2 (Reference No. 312483), “Investissement d’Avenir” (Reference No. ANR-10-EQPX-38-01) and CPER programs.

Luis Alfredo RODRÍGUEZ (Toulouse), Cristina BRAN, David REYES, Christophe GATEL, Eider BERGANZA, Manuel VÁZQUEZ, Agustina ASENJO, Etienne SNOECK
11:00 - 11:15 #6263 - MS06-OP290 Magnetic Skyrmions in an FeGe Nanostripe Revealed by in situ Electron Holography.
Magnetic Skyrmions in an FeGe Nanostripe Revealed by in situ Electron Holography.

Intense research interest in magnetic skyrmions is presently driving the development of new fundamental concepts and applications1. Magnetic skyrmions are particle-like, topologically protected swirling spin textures, in which the peripheral spins are oriented vertically, the central spins are oriented in the opposite direction and the intermediate spins rotate smoothly between these two opposite orientations, as shown in the inset to Fig. 1(a). In a range of applied magnetic fields, skyrmion lattices form in certain chiral magnets, such as B20-type magnets, in which a lack of inversion symmetry and spin-orbit coupling gives rise to the Dzyaloshinskii-Moriya interaction. The typical sizes of skyrmions are between 3 and 100 nm. For technically relevant applications, a full understanding of skyrmion formation, stability, manipulation and annihilation is required. Recent experiments have demonstrated the formation of magnetic skyrmion chains in geometrically confined nanostructures2, as shown schematically in Fig. 1(b). A critical step towards real-world device applications involves the development of an approach that can be used to controllably create, manipulate and annihilate skyrmions in magnetic nanostructures, including wire-like geometries.

Real-space imaging of complex skyrmion spin configurations using Lorentz microscopy (LM) in the transmission electron microscope (TEM) has enabled the direct observation of skyrmion lattice formation and transformations between different magnetic states with nanometre spatial resolution3. However, the finite size and the inherently weak magnetization of such magnetic nanostructures imposes great experimental challenges for LM. In particular, Fresnel fringe contrast at the specimen edge makes extremely difficult to use LM to obtain magnetic signals in samples that have lateral dimensions of below 10 nm. In contrast, off-axis electron holography (EH) in the TEM, which allows electron-optical phase images to be recorded directly with nanometre spatial resolution and high phase sensitivity, provides easier access to magnetic states in nanostructures. Digital acquisition and analysis of electron holograms and sophisticated image analysis software are then essential in studies of weak and slowly varying phase objects such as magnetic skyrmions4.

Here, we use both LM and EH to study magnetic skyrmions in a B20-type FeGe nanostripe. The use of liquid nitrogen specimen holder (Gatan model 636) allows the specimen temperature to be varied between 95 and 370 K, and the objective lens of the microscope (FEI Titan 60-300) can be used to apply magnetic fields to the specimen of 0 to 1.5 T. The aim of our study is to resolve the fine magnetic structures of geometrically confined skyrmions and to understand their formation process. Figures 2(a-b) show Lorentz images of a typical FeGe nanostripe, in which a helix to skyrmion transition occurs in response to an applied magnetic field. Figure 2(c) shows a colour-contour composite map derived from a phase image recorded using EH. The slight asymmetry of the contours results from the wedge-shaped specimen thickness profile. Artefacts associated with local changes in specimen thickness in such images can be removed from such images by separating the mean inner potential contribution from the magnetic contribution to the phase, for examples by evaluating the difference between phase images recorded at two different specimen temperatures.

References:

1 N. Nagaosa and Y. Tokura, Nat. Nanotechnol. 8, 899 (2013).

2 H. Du and et al., Nat. Commun. 6, 8504 (2015).

3 X.Z. Yu and et al., Nature 465, 901 (2010).

4 H.S. Park and et al., Nat. Nanotechnol. 9, 3 (2014).

5 Finacial support from European Research Council (ERC) Advanced Grant: IMAGINE is acknowledged. 

Zi-An LI (Duisburg, GERMANY), András KOVÁCS, Amir TAVABI, Chiming JIN, Haifeng DU, Mingliang TIAN, Michael FARLE, Rafal DUNIN-BORKOWSKI
11:15 - 11:30 #6927 - MS06-OP296 Novel spectroscopy with atomic-size aberrated electron probes in stem.
Novel spectroscopy with atomic-size aberrated electron probes in stem.

it has been theoretically argued that atomic-size electron probes with customized phase distributions can detect electron magnetic circular dichroism (EMCD) [1].  Based on this prediction we have recently shown that deliberately aberrated electron probes in scanning transmission electron microscopy (STEM) can be utilized to obtain chiral dichroic signals in materials via electron energy-loss spectroscopy (EELS) with high spatial resolution [2].

The experiments were performed in an aberration-corrected Nion UltraSTEMTM 100, equipped with a cold field emission electron source and a corrector of third and fifth order aberrations, operating at an accelerating voltage of 100kV [3].  EEL spectra were collected using a Gatan Enfina spectrometer, with 0.3 eV/channel dispersion, giving an energy resolution of 0.9 eV.  The convergence semi-angle for the incident probe and the EELS collection semi-angle were 30 mrad and 48 mrad, respectively. 

Figure 1 shows two examples of drift (affine)-corrected Z-contrast images and denoised EEL spectra that were acquired simultaneously from the room temperature C-type antiferromagnet LaMnAsO.  The data were acquired with the beam along the c-axis using a corrected electron probe (Fig. 1a to 1c) and the C34 = 15 μm aberrated probe (Figs. 1d to 1f) [2].  

 A clear EMCD signature in the EEL spectra, defined as (Mn↑ - Mn↓) presenting a change of sign in its integrated intensity between the L3 and L2 peaks, is only visible for the Mn L-edge acquired using the C34 aberrated probe. In Figs. 1e and 1f, this signal, shown in green, is positive at the L3 peak and negative at the L2 peak.  For comparison, the Mn signal in 1b and 1c, shown in grey, does not display this distinctive signature, with both positive and negative components on both peaks.

We will discuss the experimental conditions necessary to reveal the magnetic ordering of individual atomic columns and atomic-size defects in materials [4].

[1] J. Rusz, J. C. Idrobo, and S. Bhowmick, Phys. Rev. Lett. 113, (2014) p. 145501.

[2] J.C. Idrobo et al., submitted (2016).

[3] O. L. Krivanek, et al.,Ultramicroscopy 108, (2008). p. 179-195.

[4] Research supported by Oak Ridge National Laboratory’s Center for Nanophase Materials Sciences (CNMS), which is a U.S. Department of Energy (DOE), Office of Science User Facility (JCI), by the Swedish Research Council and Swedish National Infrastructure for Computing (NSC center) (JR), and by the Materials Sciences and Engineering Division, Office of Basic Energy Sciences, U.S. DOE (MAM, CC, ARL), and by UT-Battelle, LLC, under Contract No. DE-AC05-00OR22725 with the U.S. Department of Energy (CTS, RRV). 

Juan Carlos IDROBO (Oak Ridge, USA), Ján RUSZ, Jakob SPIEGELBERG, Michael A. MCGUIRE, Christopher T. SYMONS, Ranga Raju VATSAVAI, Claudia CANTONI, Andrew R LUPINI
11:30 - 11:45 #6249 - MS06-OP288 Quantifying the hole distribution in cuprates: Atomic-resolution near-edge fine-structures of the superconductor Sr3Ca11Cu24O41.
Quantifying the hole distribution in cuprates: Atomic-resolution near-edge fine-structures of the superconductor Sr3Ca11Cu24O41.

Sr14-xCaxCu24O41 is a fascinating member in the family of cuprates, not only because of its peculiar crystal structure where two distinct units, corner-shared CuO2 chains and edge-shared Cu2O3 ladders, coexist within the unit cell but also because it is the only known superconductor with a non two-dimensional CuO2 plane structure. Indeed, the superconducting state theoretically predicted by E. Dagotto and T.M. Rice [1] was first observed experimentally in Sr0.4Ca13.6Cu24O41.84 below Tc = 12 K and for pressures starting from 3 GPa [2].

            Independently of the Ca composition, Sr14-xCaxCu24O41 is an intrinsically hole-doped compound with 6 holes per formula unit, leading to an average Cu valence of +2.25. A central issue for understanding the mechanisms leading to superconductivity in this compound is therefore to measure accurately the carrier distribution among CuO2 chains and Cu2O3 ladders. This task has been undertaken shortly after the discovery of superconductivity in this system [3] but is still a matter of intense debate due to the very scattered nature of the results. For instance, depending on the technique, reported hole concentrations in the ladder layers of Sr3Ca11Cu24O41 vary from ~1 to ~4.5 holes/formula unit [4,5].

            All these results have been obtained with techniques that have a relatively poor spatial resolution ranging from several hundred nanometers to a few micrometers. In this work, we exploit the unmatched spatial-resolution of STEM-EELS to measure local hole concentration in superconducting Sr3Ca11Cu24O41 at the atomic scale and provide, for the first time, a real-space measurement where spatial separation between chains and ladders is achieved [6]. As shown by F.C. Zhang and T.M. Rice [7], in doped cuprates, the hole strongly binds to the four O atoms surrounding the central Cu through Cu3d-O2p in-plane sigma hybridization within the CuO4 plaquettes. As such, the local hole concentration can be monitored very efficiently through the O-K pre-edge structures as shown in Figure 1. These experimental results, combined with inelastic scattering calculations, demonstrate unambiguously that holes lie preferentially within the CuO2 chains of the structure.

            In summary, this work illustrates how the combination of near-edge fine-structure analyses with atomic resolution in the aberration corrected STEM can improve the understanding of the electronic properties of complex oxides, such as Cu-based superconductors [8].

 

[1] E. Dagotto and T.M. Rice, Science 271 (1996) 618.

[2] M. Uehara et al., J. Phys. Soc. Jpn. 65 (1996) 2764.

[3] T. Vuletic et al., Physics Reports 428 (2006) 169.

[4] M.-J. Huang et al.  Phys. Rev. B 88 (2013) 014520.

[5] A. Rusydi et al., Phys. Rev. B 75 (2007) 104510.

[6] M. Bugnet et al., Science Advances 2 (2016) e1501652.

[7] F.C. Zhang and T.M. Rice, Phys. Rev. B 37 (1988) 3759.

[8] The experimental work has been performed at the Canadian Centre for Electron Microscopy, a national facility supported by NSERC, the Canada Foundation for Innovation and McMaster University. Financial support by the Austrian Science Fund (FWF) under grant nr. J3732-N27 is gratefully acknowledged.

Matthieu BUGNET, Guillaume RADTKE (PARIS), Stefan LÖFFLER, Peter SCHATTSCHNEIDER, David HAWTHORN, Hanna A. DABKOWSKA, Graeme M. LUKE, George A. SAWATZKY, Gianluigi A. BOTTON
11:45 - 12:00 #6151 - MS06-OP286 Combined EELS-EDX analysis of nanoscale memristive NbOx and AlOx layers.
MS06-OP286 Combined EELS-EDX analysis of nanoscale memristive NbOx and AlOx layers.

Memristive devices – electronic components that can change their resistance depending on their history of operation – offer a new approach in various fields of digital computing. [1] This advancement in information technology is being pursued in order to satisfy the ever increasing need for computing power on the one hand and to enable the imitation of neuronal networks like the hippocampus on the other. The conductive properties in memristive devices are typically governed by atomistic effects like cation or anion movement: the change of resistance state is dependent on the nanoscopic layout. The mechanisms, however, behind a non-binary, remanent, reversible and repeatable change of electrical resistance are not well understood. In this work we present the results of dedicated transmission electron microscopy (TEM) analysis involving highly precise electron energy loss spectroscopy (EELS) methods in a complex oxide multilayer stack of only about ten nanometer total thickness. The layer sequence Nb/Al/Al2O3/NbxOy/Au (from bottom to top; Fig. 1 a) was deposited onto a Si/SiO2 substrate. The memristive properties of such junctions were recently investigated. [2]
Combined energy dispersive X-ray spectroscopy (EDX) and EELS experiments allow for analyses of both light and heavy metals as well as the reliable detection of oxygen. In contrast to the deposited layer sequence we observe an entire oxidation of the metallic Al and even a partial oxidation of the Nb bottom electrode. (Fig. 1 b) and Fig. 2) Furthermore, the results imply the abundant presence of oxygen vacancies apparent from the O K-peak in the spectrum of the amorphous Al oxide layer which acts as a tunnel barrier. (Fig. 1 c) These puzzling results will be discussed in the framework of former studies on the Nb-Al overlayer technique. [3]
Different conduction mechanisms are being discussed based on the findings and an outlook onto further research by electronic structure calculation is given. The results shed light on the fundamentals of tunnel barrier-based memristive devices which are compatible to state-of-the-art CMOS technology and were already built into integrated circuits.

Acknowledgements: The research was conducted within the DFG program FOR2093 “Memristive devices for neuronal networks”. This research has received funding from the European Union within the 7th Framework Program (FP7/2007-2013) under Grant Agreement no. 312483 (ESTEEM2).

[1] G. S. Rose, “Overview: Memristive devices, circuits and systems,” Circuits and Systems (ISCAS), Proceedings of 2010 IEEE International Symposium, Paris, 2010, 1955-1958.
[2] M. Hansen, M. Ziegler, L. Kolberg, et al. “A double barrier memristive device”. Scientific Reports. 2015, 5, 13753.
[3] J. Kwo, G. K. Wertheim, M. Gurvitch and D. N. E. Buchanan, “X‐ray photoemission spectroscopy study of surface oxidation of Nb/Al overlayer structures,” Appl. Phys. Lett. 1982, 40, 675.

Julian STROBEL (Kiel, GERMANY), Mirko HANSEN, Georg HABERFEHLNER, Gerald KOTHLEITNER, Martin ZIEGLER , Hermann KOHLSTEDT, Lorenz KIENLE
12:00 - 12:15 #6315 - MS06-OP291 Discrete spectroscopic electron tomography: using prior knowledge of reference spectra during the reconstruction.
Discrete spectroscopic electron tomography: using prior knowledge of reference spectra during the reconstruction.

A three-dimensional (3D) characterization of the morphology of nanostructures can nowadays routinely be obtained using electron tomography. [1] Nevertheless, resolving the chemical composition of complex nanostructures in 3D remains challenging and the number of studies in which electron energy loss spectroscopy (EELS) is combined with tomography is limited. [2-5] In most of these studies, two dimensional (2D) elemental maps of the object are first extracted at each tilt angle and used as an input for tomographic reconstruction. [2,3] An alternative approach is to reconstruct each energy loss separately yielding a 4D data cube where an EELS spectrum can be extracted from each 3D voxel. [4,5] During the last decade, dedicated reconstruction algorithms have been developed for HAADF-STEM tomography which use prior knowledge about the investigated sample. For example, the discrete algebraic reconstruction technique (DART) is based on the idea that a 3D HAADF-STEM reconstruction of a (nano)material only contains a limited number of grey values. [6] In this manner, several artefacts, typical to electron tomography, are mininized leading to reconstructions with a higher reliability. An additional advantage of discrete tomography is that the quantification of the final reconstruction is straightforward since the segmentation is part of the reconstruction algorithm. Here, we will extend discrete tomography to its application for spectroscopic datasets where it is assumed that the experimental spectrum of each reconstructed voxel is a linear combination of a well-known set of references spectra. 

 

To investigate the performance of discrete spectroscopic electron tomography, a phantom object is made resembling a Ce4+ nanoparticle with a reduced Ce3+ edge as presented in Figure 1a. A tilt series of projected spectrum images are simulated and different amounts of Poisson noise are applied to the projection data. These datasets are used as input for two conventional reconstruction approaches and the discrete spectroscopic reconstruction technique. In the first method, elemental maps are first extracted which are used to reconstruct the individual chemical elements. The second method first reconstructs all energy losses yielding a complete 4D dataset. The reconstructions of the different elements are then obtained using a spectrum fitting procedure. The average reconstruction error as a function of the signal to noise ratio (SNR) is displayed in Figure 1b. This graph indicates that  discrete spectroscopic electron tomography, displayed in red, provides superior results especially for datasets with a relatively low SNR. Therefore, it is well suited for the 3D reconstruction of small dopants in nanoparticles typically having a low SNR in the projected spectrum images.

 

Next, we investigated the spatial distribution of Fe dopants in Fe:Ceria nanoparticles. During the tomographic reconstruction, reference spectra for Fe2+, Ce3+ and Ce4+ are used as prior knowledge. Visualizations of the final reconstructions are presented in Figure 2. As indicated by the white arrows, we can observe that the presence of the Fe2+ dopants is correlated with a reduction of the Ce atoms from Ce4+ towards Ce3+. This indicates that both the Ceria nanoparticle and the Fe dopants are reduced by the generation of oxygen vacancies. In addition, from the comparison of the slices through the HAADF-STEM reconstruction and the Fe2+ reconstruction (Figure 2f), it can be observed that most of the Fe dopants are located near the voids of the nanoparticle.

 

[1] P.A. Midgley, R.E. Dunin-Borkowski, Nat. Mater. 8 (2009) 271-280

[2] L. Yedra, et al., Ultramicroscopy 122 (2012) 12-18

[3] O. Nicoletti, et al., Nature 502 (2013) 80-+

[4] G. Haberfehlner, et al., Nanoscale 6 (2014) 14563-14569

[5] B. Goris, et al., ACS Nano 8 (2014) 10878-10884

[6] K.J. Batenburg, et al., Ultramicroscopy 109 (2009) 730-740

[7] The authors acknowledge funding from the Research Foundation Flanders (project number G038116N and a post-doctoral grant to B.G.). S.B. acknowledges the European Research Council, ERC grant N°335078 – Colouratom.

Bart GORIS (Antwerp, BELGIUM), Maria MELEDINA, Stuart TURNER, Zhichao ZHONG, Joost BATENBURG, Gustaaf VAN TENDELOO, Sara BALS
12:15 - 12:30 #6638 - MS06-OP292 Resolving the atomic and electronic structures of bismuth iron garnet thin films.
Resolving the atomic and electronic structures of bismuth iron garnet thin films.

Bismuth iron garnet (Bi3Fe5O12- BIG) presents large interests in both fundamental studies and applications thanks to a plethora of fascintating physical properties. Among them, BIG is ferrimagnetic with a relatively high magnetization of 1600 G at 300 K and presents magnetic ordering temperature from 650 K to 700 K [1]. One of its most useful properties is the giant Faraday rotation (~105 deg/cm at 550 nm) making it potential for magneto-optical recording and the fabrication of non-reciprocal magneto-optical devices based on magnetophotonic crystals [2]. More recently, a strong magneto-electric coupling at room temperature has been reported in BIG thin films opening new perspectives for an electric control of the magnetization [3].

Contrary to its parent structures, e.g. the well-known yttrium iron garnet (YIG), this material can only be elaborated in thin film form using non-equilibrium growth techniques. BIG films were grown onto distinct isostructural Y3Al5O12 (YAG) and substituted-Gd3Ga5O12(SGGG) substrates; each is supposed to induce a compressive epitaxial strain depending on the lattice mismatch, i.e. approx. of 5% and 1%, respectively. The interplay between strain engineering and control of its promising magnetic and transport properties is of main interest in these systems, however no bulk reference is available yet for detailed structural and magnetic studies. Hence resolving the atomic and electronic structures of BIG thin films remains a key challenge to understand better their structure-property relationship. Here we use advanced spectromicroscopy studies combining a Cs-corrected scanning transmission electron microscope (STEM), the NION UltraSTEM200, with high-energy resolved electron energy-loss spectroscopy (EELS) experiments to characterize these BIG thin films, from their structural aspect to their elemental and spectroscopic properties.

Peculiar relaxation mechanisms are highlighted in both BIG/substrate systems by Geometric Phase Analysis (GPA) based on low-magnified high-resolution HAADF-STEM images. All the films are epitaxially grown and relaxed from thicknesses above a few tens of nanometers. Combined GPA studies and atomically-resolved HAADF-STEM images at the interfaces reveal how BIG can accommodate different lattice mismatches through a variety of growth mechanisms. Fig.1 gathers the dominant scenarii observed in BIG thin films with the typical “cube-on-cube” growth at the BIG/SGGG interface or the more complexed Vernier misfit alternating with tilted grains presenting a network of dislocations in the BIG/YAG system. The evolution of the local in-plane and out-of-plane lattice parameters confirms a rather strong compressive strain of the BIG film grown onto SSGG occurring at the first few interfacial unit cells. Whereas an immediate relaxation is observed at the BIG/YAG interface with a relatively uniform evolution of both local in-plane and out-of-plane parameters revealing a quasi-absence of strain thanks to the particular coexistence of tilted grains and well-relaxed Vernier misfit.

Finally, all BIG films seem to preserve the garnet cubic structure (Fig. 2 a). From a spectroscopic study, no cation interdiffusion is evidenced when probing the film/substrate interface down to the scale of the atomic columns, and only the signature of Fe3+ valence state is detected confirming the absence of oxygen deficiency. Besides, the evolution of the O-K near-edge fine-structures through the BIG/YAG interface enables us to figure out directly possible electronic reconstruction at the interface by probing the Fe 3d - O 2p hybridization (Fig. 2b).

[1] E. Popova et al., J. Magn. Magn. Mater. 335, 139 (2013).

[2] M. Deb, et al. J. Phys. D: Appl. Phys. 45, 455001 (2012)

[3] E. Popova et al. submitted

Laura BOCHER (Orsay), Adrien TEURTRIE, Elena POPOVA, Bénédicte WAROT-FONROSE, Niels KELLER, Odile STÉPHAN, Alexandre GLOTER

10:15-12:30
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MS5-II
MS5: Energy-related materials
SLOT II - Battery materials

MS5: Energy-related materials
SLOT II - Battery materials

Chairmen: Wolfgang JÄGER (Kiel, GERMANY), Joachim MAYER (Aachen, GERMANY), Philippe MOREAU (Associate Professor) (Nantes, FRANCE)
10:15 - 10:45 #6672 - MS05-S78 Atomic Resolution STEM and Spectroscopic Characterization of Battery Related Materials.
Atomic Resolution STEM and Spectroscopic Characterization of Battery Related Materials.

The properties of lithium battery strongly depend on the diffusion of lithium ions during charge/discharge process. Since this behavior determines the stability, lifetime and reliability, direct visualization of Li site is required to understand the mechanism of the diffusion of lithium ions. Annular bright field (ABF) scanning transmission electron microscopy (STEM) is useful imaging technique to directly observe the both light and heavy element columns [1]. In this technique, an annular detector is located within the bright-field (direct-scattered) region, and the columns display absorptive-type contrast. Figure 1 shows ABF STEM images observed from [001] of (a) olivine LixFePO4 and (b)delithiated olivine (FePO4). It can be seen that Li column contrast appears in LixFePO4, but disappears in FePO4. Fig.1 (c) (d) shows HAADF STEM  images observed from the same region of (a) and (b), indicating that the cation frame work columns are almost the same before and after delithiation [2]. In this study, light elements in several lithium battery related materials are directly observed by ABF STEM, and the mechanism of lithiation/delithiation is discussed based on the observation results.

The properties of thin-film batteries is influenced by the atomic structures of the embedded interfaces, such as electrode/electrolyte and electrode/current-collector interfaces, as well as the grain and domain boundaries. Detailed analyses of these interface structures, which provides insights into formation mechanisms of the interfaces and the effects of microstructure on electrochemical properties, is essential for understanding the mechanism of lithiation /delithiation and for obtaining the guideline to design the thin film devices. In this study, the epitaxial growth mechanism of a typical cathodic LiMn2O4 thin film is investigated by exploring the detailed structural and compositional variations in the vicinity of the film/substrate interfaces. STEM observation shows the LiMn2O4 film forms an atomically flat and coherent heterointerface with the Au(111) substrate, but that the crystal lattice is tetragonally distorted with a measurable compositional gradient from the interface to the crystal bulk [3]. The growth mechanism is interpreted from the chemical and physicomechanical effects, which is related to the complex interaction between the internal Jahn-Teller distortions induced by oxygen non-stoichiometry and the lattice misfit strain.

In addition, the microstructures for La2/3-xLi3xTiO3 (LLTO) and La(1-x)/3LixNbO3 (LLNO) electrolytes are characterized by ABF STEM. It was found that the unique structures of the domain boundaries (DBs) in LLTO affect the Li-ion mobility and ionic conductivity. DBs in LLTO are consisted of two types: frequently occurring 90° rotation DBs and a much less common antiphase-type boundary [4]. The 90° DBs are found to have coherent interfaces consisting of interconnected steps that share La sites, with occupancies of La sites higher than in the domain interiors. The DBs show different degrees of lattice mismatch strain depending on Li content. The lattice strain and resultant Li and O vacancies and the high La occupancy at DBs are considered to be the reason for lower interdomain Li-ion mobility, which has a deleterious effect on the overall Li-ion conductivity. LLNO is found to have complex modulated crystal structures with partially ordered distributions of A cations and vacancies. This involves a long-range layer-ordering of A cations into alternating La/Li-rich and La/Li-free layers parallel to (001)p, and a short-range sinusoidal columnar ordering of A cations within the La-rich layers.

 References

[1] S. D. Findlay et al., Appl. Phys. Lett., 95, 19191 (2009); [2] A. Nakamura et al., Chem. Mater., 26, 6178 (2014) ; [3] X.Gao, et.al., Adv.Mater.Inter., 1400143 (2014); [4] X. Gao, et. al., J. Mater. Chem. A, 3,3351 (2015). [5] This research is supported by Japan Fine Ceramics Center and Toyota Motor Co.. A part of this work was also supported by the Research and Development Initiative for Scientific Innovation of New Generation Batteries (RISING) project from the New Energy and Industrial Technology Development Organization (NEDO), Japan.

Yuichi IKUHARA (tokyo, JAPAN)
Invited
10:45 - 11:00 #6875 - MS05-OP276 In situ study of the degradation phenomena induced by lithiation/delithiation cycle of a composite Si-based anode by the mean of X-ray tomography.
In situ study of the degradation phenomena induced by lithiation/delithiation cycle of a composite Si-based anode by the mean of X-ray tomography.

In the context of increasing energy density of lithium-ion batteries, silicon is of high interest with his high theoretical gravimetric capacity, ten times higher than the commonly used carbon. However the use of silicon is faced to huge hurdles such as poor life time of those electrodes and not sufficient sustainability versus high current density (commonly used in EVs and HEVs).

The poor cycle life of Si-based electrodes is mainly due to their large volume variation upon cycling, inducing electrical disconnections and instability of the solid electrolyte interface (SEI). The study of the morphological variation of Si-based electrodes upon cycling is thus highly relevant to evaluate their degradation and to optimize their formulation and architecture. However, this is challenging considering their complex three-dimensional structure and their major evolution with cycling. Furthermore, samples are fragile and reactive and therefore difficult to prepare for bulk observations. In this context, X-ray tomography appears as an effective non-destructive and 3D observation tool.

 

In this communication, in-situ synchrotron  X-ray tomography analyses are performed on Si-based electrodes prepared from a pH3 buffered slurry of ball-milled Si powder + carbon black + carboxymethylcellulose (CMC) (80/12/8) loaded into a carbon paper (AvCarb EP40) by impregnation, in order to get a clear view of the 3D architecture of the electrode with cycling. From the initial state, represented in Fig. 1, to fully lithiated and then fully delithiated state, X-ray scans were performed each thirty minutes to continuously follow the morphological evolution of the electrode structure.

After an appropriate image reconstruction and segmentation procedure, phase identification has been achieved. Moreover the separation of a void porosity and an electrolyte phase was possible and the quantifying of the pore size distribution evolution with cycling, as shown in Fig. 2.

Also key morphological parameters of these Si-based electrodes and their evolution with cycling are determined, such as the electrode thickness and volume fraction of the pores as shown in Fig. 3. Those results may greatly enlighten the understanding of degradation phenomena in the Si-based electrodes and help develop new composite electrodes formulation for sustainable applications.

Victor VANPEENE (Lyon), Eric MAIRE, Aurélien ETIEMBLE, Lionel ROUÉ, Anne BONNIN
11:00 - 11:15 #5315 - MS05-OP263 Quantitative electron diffraction tomography for the structure solution of cathode materials for Li-ion batteries.
Quantitative electron diffraction tomography for the structure solution of cathode materials for Li-ion batteries.

Quantitative electron diffraction tomography (EDT) is a perfect tool for studying electrode materials for Li-ion batteries due to its ability to obtain diffraction data from submicron crystals (typically 100-200 nm), sensitivity to “light” atoms as Li and reduced dynamical effects (enhanced by the intrinsic property of the electrode materials to comprise only lightest possible elements to maximize the capacity). Here we demonstrate applications of EDT to the crystal structure analysis of polyanion cathodes, including location and occupancy refinement of the Li positions.

We applied EDT  for monitoring the deintercalation process in LiMn0.5Fe0.5PO4 (LMFP) olivine [1]. We investigated a pristine and two delithiated phases (mid (3.7 V vs. Li/Li+) and fully-charged (4.2 V), since upon charge two first-order phase transformations take place). All structures keep the Pnma space group and unit cell metrics and no sign of local ordering was found. The difference Fourier maps of the electrostatic potential calculated from EDT data, allow to localize the Li atoms (Fig 1). The refined occupancies of the Li positions for all samples are in excellent agreement with the Li contents measured from the E-x dependence obtained with galvanostatic cycling. Regarding the role of Jahn-Teller distortion due to Mn3+ in the delithiation mechanism, this effect is not of cooperative nature, since the octahedral distortions in pristine and fully-charged samples vary insignificantly (5%).

EDT sheds light on the family of (Li,A)-ion cathode materials that are obtained by subsequent chemical or electrochemical exchange of Na/K by Li. This allows obtaining new polymorphs that are not attainable by direct synthesis due to thermodynamic reasons. We synthesize new KVPO4F fluorophosphates with KTiOPO4 (KTP) structure, having a 3D system of continuous spatial cavities and two potassium positions [2]. Upon charge up to 5.0 V vs Li/Li+ the K1 site becomes empty whereas residual potassium (17%) resides in the K2 site (Fig. 2). At the same time, the structure changes from noncentrosymmetric (Pna21) to centrosymmetric (Pnan). After lithiation, the K2 sites are shared by K and Li but K1 still remains empty. Li occupies a new Li3 position having [4 + 2] coordination with four short 1.99−2.14 Å Li3−O bonds. Both Li sites reside in the channels along the b axis and form a row of Li atoms.

The discussed exapmles show that EDT is a relible technique for the crystal structure solution and refinement. It can be used as a routine method for the study of cathode materials, since no special sample preparation is required. Furthermore, EDT provides valuable information of the lithium atom positions and occupancies.

Acknowledgements

J. Hadermann, O. M. Karakulina and A. M. Abakumov acknowledge support from FWO under grant G040116N. S.S. Fedotov acknowledges support from RFBR (grant 16-33-00211 mol_a).

[1]      O.A. Drozhzhin, V.D. Sumanov, O.M. Karakulina, A.M. Abakumov, J. Hadermann, A.N. Baranov, K.J. Stevenson, E. V Antipov, Electrochim. Acta 191 (2016) 149.

[2]      S.S. Fedotov, N.R. Khasanova, A.S. Samarin, O.A. Drozhzhin, D. Batuk, O.M. Karakulina, J. Hadermann, A.M. Abakumov, E. V. Antipov, Chem. Mater. 28 (2016) 411.

Olesia KARAKULINA (Antwerp, BELGIUM), Stanislav FEDOTOV, Vasiliy SUMANOV, Oleg DROZHZHIN, Nellie KHASANOVA, Evgeny ANTIPOV, Artem ABAKUMOV, Joke HADERMANN
11:15 - 11:30 #5556 - MS05-OP264 In situ scanning and transmission electron microscopy experiments developed at the University of Picardy: a versatile approach optimized for the study of lithium-ion batteries and extended to the observation of plant tissues.
In situ scanning and transmission electron microscopy experiments developed at the University of Picardy: a versatile approach optimized for the study of lithium-ion batteries and extended to the observation of plant tissues.

Over the past fifteen years, the Platform of Electron Microscopy at the University of Picardy Jules Verne (PME UPJV) has been developing in situ scanning electron microscopy techniques to successfully observe polymer [1] and solid or liquid electrolyte lithium-ion batteries cycling in the scanning electron microscope (figure 1). In situ Electron Microscopy techniques allow to spot the slightest textural, chemical or structural modification of lithium-ion electrodes, interfaces and active materials.  Consequently, such innovative studies help electrochemists to design batteries with better performance through the selection/synthesis of the appropriate electrode materials and the control of the electrode/electrolyte interfaces upon cycling.

However, considering the 'resolution limit' of the SEM, electrochemists working on lithium-ion batteries tend towards in situ transmission electron microscopy to fulfill their ambition of atomic-scale observations of electrochemical cycling. Nevertheless, there are several technological bottlenecks in using such techniques. For instance restrictions imposed by the limited sample size and thickness (less than 100nm) and the requirement of high vacuum necessitate the use of adapted  strategies (all-solid-state batteries, ionic liquid as electrolyte, STM or liquid bias TEM holder)[2,3,4]. To address these issues, we undertook two different approaches:  cycling all-solid-state microbatteries with a Nanofactory STM holder [5] (figure 2) and cycling liquid electrolyte batteries with a Protochips liquid/bias Holder.

We are extending now the application of these developed methods at PME UPJV to perform in situ analysis of samples from another branch of natural science: Plant Biology (from seeds to biorefinery). The in situ electron microscopy analyses will be performed to characterize the role of plant specialized cell layers (figure 3) and lignocellulosic biopolymers and examine their modification as a result of seed germination, enzymatic modifications, micromechanical tests as well as plant pathogen interactions.

We gratefully acknowledge the Region of Picardy, The French National Research Agency, ALISTORE-ERI and the European Social Fund for their financial support.

References

[1]          Orsini et al., Int. J. Inorg. Mater. 2:6 (2000) 701.

[2]          Baer et al., J. Mater. Res. 25:8 (2010) 1541.

[3]          Huang et al., Science. 330:6010 (2010) 1515.

[4]          Yamamoto et al., Angew. Chem. Int. Ed. 49:26 (2010) 4414.

[5]          Brazier et al., Chem. Mat. 20:6 (2008) 2352.

Loic DUPONT (amiens cedex), Arnaud DEMORTIERE, Carine DAVOISNE, Arash JAMALI, Walid DACHRAOUI, Mattia GIANNINI, Fabien MIART, Damien MC GROUTHER, Rick BRYDSON
11:30 - 11:45 #6163 - MS05-OP269 Low-loss STEM-EELS analysis of beam-sensitive lithium-ion negative electrodes.
Low-loss STEM-EELS analysis of beam-sensitive lithium-ion negative electrodes.

Silicon represents one of the most promising anode materials for next generation lithium-ion batteries. However its colossal volume expansion (up to 300%) upon electrochemical reaction with lithium repeatedly exposes fresh surfaces to electrolyte solvent oxidation1-2. This leads to very high irreversible capacities, compounded by the fact that parts of the silicon-based electrodes are progressively disconnected from both electrical and ionic transport networks as the solid electrolyte interface (SEI) accumulates. Deeper insight into these degradation phenomena is critical to engineer adequate electrodes and/or electrolytes.

 

Characterization of these electrodes has so far mostly focused on either bulk or surface analysis, both lacking spatial resolution. Little is known about the SEI’s morphology in silicon nanoparticles (SiNPs) aggregates that make the electrode, or about the evolution of these nanoparticles themselves with cycling. Attempts to characterize this system through electron microscopy have been severely limited by the radiolysis and sputtering damage, respectively, undergone by the SEI and lithium-silicon alloys (LixSi).

 

In this work we demonstrate the possibility to map major SEI and electrode components such as lithium carbonate (Li2CO3), lithium fluoride (LiF) and lithium oxide (Li2O) as well as quantifying lithium-silicon alloys compositions4 and Si crystallinity from a single dataset by combining scanning transmission electron microscopy and low-loss electron energy loss spectroscopy5 (STEM-EELS) (fig. 1). The low-loss part of the EEL spectrum is considerably more intense than its high energy counterpart and contains both the Li K-edge and plasmons. Fine tuning of the experimental parameters allows us to acquire low-loss spectrum images with good signal-noise ratios within timeframes compatible with minimal sample degradation. Plasmons can then either be used as unique molecular signatures for the SEI, or directly for quantification in the case of LixSi compounds (fig. 2 inset). This can yield unique insight into electrode degradation phenomena through careful data processing (MLLS, Drude model fit...). Large spectrum images can be acquired within short timeframes (~10 ms/voxel), making this method a powerful and practical diagnostics tool for battery electrodes and other beam-sensitive nanostructured systems.

 

Results on electrodes disassembled from full cells at their 1st, 10th and 100th charge and discharge, with a limited capacity of 1200 mAh/g, shed light on the SEI’s deposition mechanism and morphological as well as chemical evolution along cycling for different electrolytes. Strong correlations were observed between the SEI's local chemistry and our nanoparticles cycling performance (fig. 2). Lithiation was also observed to proceed preferentially along grain boundaries, resulting in different behaviours between mono- and polycrystalline silicon powders.

 

 

(1)          Delpuech et al. ChemSusChem 2016, 9, 1-9

(2)          Dupré, Boniface et al. Chem. Mater 2016 (accepted)

(3)          Danet et al. Phys. Chem. Chem. Phys. 201012 (1), 220–226.

(4)          Yakovlev et al. M. Micron 200839 (6), 734–740.

(5)          Egerton, R. F. Reports Prog. Phys. 200972 (1), 016502. 

Maxime BONIFACE (Grenoble), Lucille QUAZUGUEL, Philippe MOREAU, Florent BOUCHER, Dominique GUYOMARD, Pascale BAYLE-GUILLEMAUD
11:45 - 12:00 #4929 - MS05-OP260 Comparison of energy filtered TEM spectra image and automated crystal orientation mapping in LiFePO4/FePO4 phase mapping.
Comparison of energy filtered TEM spectra image and automated crystal orientation mapping in LiFePO4/FePO4 phase mapping.

Lithium iron phosphate (LiFePO4, LFP) is one of the most promising cathode materials for the next generation of Li ion batteries and attracts great attentions. Experimental mapping the lithiated (LFP) and delithiated phase (FePO4, FP) at nanoscale resolution provides knowledge on the microscopic mechanism of the reaction processes during electrical cycling, which is crucial to improve the limits of this material. Versatile scanning / transmission electron microscopy (S/TEM) techniques, due to the advantage of intrinsic imaging ability by the electron optics, have attracted extreme interest in high resolution phase mapping. The methods are generally sorted into two kinds: one is based on electron energy loss spectroscopy (EELS), such as energy filtered TEM (EFTEM) [1], relying on the chemical information in the energy spectra; the other is automated crystal orientation mapping (ACOM) [2], originally designed for orientation analysis of nanocrystalline materials [3], relying on the crystallographic information recorded in diffraction patterns. However, so far, there is no strongly convincing evidence indicating the consistency between the chemical and the crystallographic information in the phase map, because of lacking comparison of the results between the two kind methods.

 

In this work, we applied both EFTEM and ACOM methods to the same part of a sample (half lithiated) for comparison. Maps obtained by ACOM and EFTEM of Fe-L3,2 (figure 1 a, b) show excellent agreements with each other. It proves the reliability of both methods, i.e. the consistence of the chemical and crystallographic information for the LFP/FP system. Furthermore, we demonstrate that the properties of the LFP/FP interfaces can be further characterized from the crystallographic data obtained by ACOM: on average 1.4 ° misorientation was observed at all interfaces (figure 1 d, e), and these interfaces have a preferred orientation with the normal close to the a-axis (100), but slightly deviated towards the c-axis (001) (figure 1 f, g) in agreement with [4]. Further attention is drawn to the low energy loss regime for EFTEM analysis. Figure 2 a, c and e respectively shows a map measured from the Li-K & Fe-M edges (figure 2 b), and mapping of  the dielectric function (figure 2 d) and the volume plasmon center (figure 2 f). Finally, a comprehensive comparison of all methods is given in terms of information contents, dose level, acquisition time and signal quality. The latter three are crucial for the design of in-situ experiments [5]. 

 

References:

[1] JD Sugar et al, J Power Sources 246 (2014), p512.

[2] G Brunetti et al, Chem Mater 23 (2011), p4515.

[3]  EF Rauch et al, Zeitschrift für Krist 225 (2010), p103.

[4]  M Welland et al, ACS Nano (2015), p9757.

[5]  The authors acknowledge funding from Hi-C project. Dr. Di Wang is thanked for discussions.

Xiaoke MU (Eggenstein-Leopoldshafen, GERMANY), Aaron KOBLER, Di WANG, Venkata Sai Kiran CHAKRAVADHANULA, Sabine SCHLABACH, Dorothee-Vinga SZABO, Paul NORBY, Christian KÜBEL
12:00 - 12:15 #6030 - MS05-OP268 Growth and degradation of advanced octahedral Pt-alloy nanoparticle catalysts for fuel cells.
Growth and degradation of advanced octahedral Pt-alloy nanoparticle catalysts for fuel cells.

Octahedral Pt-Ni nanoparticles are highly attractive as fuel-cell catalysts due to their extraordinarily high activity for the oxygen-reduction-reaction (ORR). A deep understanding of their atomic-scale structure, degradation and formation is a prerequisite for their use as rationally designed nanoparticle catalysts with high activity and long-term stability.

Here we present an extensive microstructural study of the growth and degradation behavior of various octahedral Pt-alloy nanoparticles using in situ transmission electron microscopy (TEM) and Cs-corrected high-angle annular dark-field scanning transmission electron microscopy (HAADF-STEM) combined with electron energy-loss spectroscopy (EELS) and energy-dispersive X-ray spectroscopy (EDX). We show that octahedral nanoparticles often show compositional anisotropy with Ni-rich {111} facets leading to complex structural degradation during ORR electrocatalysis. The Ni-rich {111} facets are preferentially etched, resulting in the formation of first concave octahedra and then Pt-rich skeletons that have less active facets (Figure 1)[1]. Furthermore, we reveal element-specific anisotropic growth as the reason for the compositional anisotropy and the limited stability. During the solvothermal synthesis, a Pt-rich nucleus evolves into precursor nanohexapods, followed by the slower step-induced deposition of Ni on the concave hexapod surface, to form octahedral facets (Figure 2)[2]. While the growth of Pt-rich hexapod is a ligand-controlled kinetic process, the step-induced deposition of the Ni-rich phase at the concave surface resembles a thermodynamically controlled process accomplished in much longer time. In order to tune the atomic-scale microstructure of the octahedra for long-term stability, we illustrate the effect of varying the growth conditions on morphology and compositional segregation by producing trimetallic PtNiCo nanooctahedra and comparing “one-step” and newly-developed "two-step" synthesis routes [3]. Furthermore we demonstrate how Pt atom surface diffusion may produce a protective Pt surface layer on top of the Ni-rich facets, resulting in advanced and more stable octahedral catalysts. Figure 3 shows a sequence of structural changes taking place on an octahedral nanoparticle during in situ heating up to 800°C using a MEMS chip heating holder (DENSsolutions, Delft, NL). It can be observed that Pt-rich corner atoms diffuse and subsequently fill the concave Ni-rich {111} facets, forming perfectly octahedral nanoparticles with flat Pt-rich {111} surfaces (Figure 3) [4].

 

[1]    Cui CH, Gan L, Heggen M, Rudi S, Strasser P, Nature Materials 2013; 12: 765.

[2]    Gan L, Cui CH, Heggen M, Dionigi F, Rudi S, Strasser P, Science 2014; 346: 1502.

[3]    Arán-Ais RM, Dionigi F, Merzdorf T, Gocyla M, Heggen M, Dunin-Borkowski RE, Gliech M, Solla-Gullón J, Herrero E, Feliu JM, Strasser P, Nano Letters 2015; 15: 7473-7480.

[4]    Gan L, Heggen M, Cui CH, Strasser P, ACS Catalysis 2016; 6: 692.

Marc HEGGEN (Jülich, GERMANY), Martin GOCYLA, Lin GAN, Peter STRASSER, Rafal DUNIN-BORKOWSKI
12:15 - 12:30 #6591 - MS05-OP272 Degradation of (La,Sr)(Co,Fe)O3-δ SOFC Cathodes at the Nanometre Scale and Below.
Degradation of (La,Sr)(Co,Fe)O3-δ SOFC Cathodes at the Nanometre Scale and Below.

     For developing solid oxide fuel cells (SOFCs) operating at intermediate temperatures, metallic materials have become a preferential choice for the interconnect due to their low cost and excellent physical and chemical properties. However the presence of chromium in all commonly used metallic alloys has been found to cause poisoning of the cathode leading to rapid electrochemical performance degradation of the cathodes including one of the most promising (La,Sr)(Co,Fe)O3-δ (LSCF) perovskite oxides [1-3]. Despite the extensive research on the chromium deposition and poisoning processes, careful microstructural studies, especially at the nanoscale, are rare, which can provide valuable information for the fundamental understanding of the Cr poisoning mechanisms required for developing Cr tolerant cathode materials.  

     In this paper, we examine the Cr poisoning mechanisms in LSCF materials by correlating the bulk electrochemical properties of the cell with their structural and chemical change at multi-scales down to the nanometer level. Cells with LSCF cathodes were prepared, and the effect of Cr poisoning on the electrochemical behavior of the cell was assessed by impedance spectroscopy. The change in nano/microstructure and chemistry due to poisoning were studied in parallel by a combination of several advanced electron microscopy techniques including focus ion beam (FIB) tomography, high resolution (scanning) transmission electron microscopy ((s)TEM) and analytical STEM. Our results show that Cr poisoned samples exhibit multiscale changes especially at the nanoscle including formation of nanometer size Cr rich phases (Figure 1), Cr segregation at LSCF grain boundaries (Figure 2), alternation of local LSCF stoichiometry and structure (Figure 3), and change of valence state of the B site elements. These observed nanoscale changes are consistent with the impedance data measured from the same samples that shows the reduction of both oxygen surface reaction rate and oxygen diffusion  by 1-2 orders of magnitude after Cr poisoning. The work revealed critical degradation mechanisms effective at the nano to atomic scale and provide new insight for the development of future poisoning resistant electrode materials not only for SOFCs but also for other devices such as solid oxide electrolysis cells.

References

[1]       M.C. Tucker, H. Kurokawa, C.P. Jacobson, L.C. De Jonghe, S.J. Visco, J. Power Sources 160 (2006) (1) 130.

[2]       S.P. Jiang, X.B. Chen, Int. J. Hydrog. Energy 39 (2014) (1) 505.

[3]       S.N. Lee, A. Atkinson, J.A. Kilner, J. Electrochem. Soc. 160 (2013) (6) F629.

Ni NA (london, UK), Cooper SAMUEL , Skinner STEPHEN

10:15-12:30
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IM10-II
IM10: Correlative microscopy
SLOT II

IM10: Correlative microscopy
SLOT II

Chairmen: Yannick SCHWAB (Heidelberg, GERMANY), Paul VERKADE (Bristol, UK)
10:15 - 10:45 #8318 - IM10-S60 Next Gen CLEM: super-accurate correlation and intelligent image acquisition.
Next Gen CLEM: super-accurate correlation and intelligent image acquisition.

Correlative light and electron microscopy (CLEM) combines the benefits of fluorescence and electron imaging, revealing protein localisation against the backdrop of cellular architecture. CLEM is usually performed by growing cells on gridded coverslips, imaging the cells (live or fixed) using confocal microscopy, preparing the cells for electron microscopy, relocating the cell position and plane for electron imaging of the fluorescent structure, selecting and modeling the 3D data in both modalities, and overlaying the two datasets to identify the structure of interest. This process usually requires collaboration with expert electron microscopists, and has sufficient steps and complexity to deter many researchers. Nevertheless, for those who commit to the process, there are rich rewards in the understanding of biological processes. Our recent work has focused on improving the speed, accuracy and accessibility of CLEM.

 

During this development work, it became clear that the technical challenges associated with CLEM are exaggerated when working in 3D. To increase protein localisation precision, we developed an ‘In-Resin Fluorescence’ (IRF) protocol that preserves the activity of GFP and related fluorophores in resin-embedded cells and tissues. The sample preparation is relatively fast, and also introduces electron contrast so that cell structure can be visualised in the electron microscope. Once the resin blocks have been cut into ultrathin sections, out-of-plane fluorescence is removed resulting in physical ‘super-resolution’ light microscopy in the axial direction, which increases the accuracy of the LM-EM overlays. Localisation precision is further increased by imaging the IRF sections in vacuo in the next generation of commercial integrated light and electron microscopes (ILEM). We were able to further improve accuracy by developing integrated super-resolution light and electron microscopy, using the novel and remarkable blinking property of GFP and YFP in-resin in vacuo, and implementing automated 3D imaging for 3D functional and structural analysis of whole cells and tissues at the nanoscale.

 

With the advent of dual fluorescence-electron samples comes the challenge of locating and following  fluorescent cells during sample preparation and automated 3D EM image acquisition. We present a new locator tool – the miniature light microscope (miniLM), designed to integrate with the ultramicrotome to locate cells during trimming and sectioning, and an even smaller version that fits into the extremely tight space of the 3D SEM vacuum chamber for on-the-fly tracking of fluorescent cells during long automated imaging runs.

Lucy COLLINSON (Londres, UK)
Invited
10:45 - 11:00 #6928 - IM10-OP175 Super-resolution fluorescence microscopy of cryo-immobilized samples.
Super-resolution fluorescence microscopy of cryo-immobilized samples.

Correlative light and electron microscopy (CLEM) benefits greatly from the development of super-resolution fluorescence microscopy. With a resolution down to the 10 nm range it enables to bridge the large resolution gap between the two different microscopy techniques. However, equally important as the achievable resolution of the imaging system is the preservation of the biological structures during the sample preparation process. Imaging undisturbed structures in living cells remains very challenging for super-resolution fluorescence microscopy with its relatively long acquisition times. Typically, chemical fixation is used to immobilize the sample for achieving best technical results, but unfortunately this is associated with structural changes in the sample, especially at a level below the diffraction limit of light [1,2].

Cryo-immobilization offers a preferable alternative. Here, fast freezing techniques enable vitrification of the sample and preserve the structures in a near-native state. Although cryo-immobilization has been established as a routine technique in the fields of electron and X-ray microscopy, it was long unclear whether super-resolution fluorescence microscopy could be performed under cryo-conditions to image vitrified samples [3].

Recently, we demonstrated on the single molecule level that photo-switching of fluorescent proteins is possible under cryo-conditions and suitable for the super-resolution method of single molecule localization microscopy (SMLM) [4]. Chang et al. and Liu et al. have shown the correlation of cryo-SMLM with electron cryo-microscopy [5,6]. Here, we present cryo-SMLM of vitrified biological samples and its prospects for cryo-CLEM. We demonstrate that a resolution improvement of up to 5x compared to conventional fluorescence cryo-microscopy is possible.

Super-resolution fluorescence cryo-microscopy offers the possibility to image fluorescently labelled biological samples with diffraction-unlimited resolution, immobilized, but with structural preservation in a near-native state. This is not only helping to bridge the large resolution gap in cryo-CLEM, but might also offer an alternative to the dilemma in conventional super-resolution imaging, where one has to choose between the limited temporal resolution in case of living cells or the disadvantages of chemical fixation.

[1] Bleck et al., 2010, J. Microsc.

[2] Weinhausen et al., 2014, Phys. Rev. Lett.

[3] Kaufmann et al., 2014, Curr. Opin. Chem. Biol.

[4] Kaufmann et al., 2014, Nano Lett.

[5] Chang et al., 2014, Nature Meth.

[6] Liu et al., 2015, Sci. Rep.

Rainer KAUFMANN (Oxford, UK), Christoph HAGEN, Kay GRÜNEWALD
11:00 - 11:15 #6135 - IM10-OP168 Electron-beam induced fluorescence superresolution with 100nm resolution in CLEM on labelled tissue sections.
Electron-beam induced fluorescence superresolution with 100nm resolution in CLEM on labelled tissue sections.

We present a novel optical superresolution (SR) technique using integrated correlative light and electron microscopy. Recent advances in SR techniques has revolutionized the field of optical microscopy by achieving image resolutions well below the diffraction limit, the fundamental resolution limit of traditional optical microscopy. Current SR methods involve stochastic techniques, beam-shaping in combination with confocal scanning, external control over excited state relaxation pathways, and/or structured illumination [1]. Correlation of SR data with ultrastructural images obtained with electron microscopy (EM) has been demonstrated [2], but requirements for SR microscopy are often in conflict with those for EM. Moreover, the optical localization accuracy in the correlation image may be severely compromised compared to the SR resolution by the additional error introduced in aligning the separate SR and EM images. Here, we demonstrate a novel approach for correlative SR-EM using a focused electron beam to locally modify the fluorescence signal of fluorophores, and detecting the change in fluorescence intensity with a wide-field epi-fluorescence microscope.

 

We use an integrated light-electron microscope [3] that is used for correlative light and electron microscopy (CLEM) [2]. The integrated light microscope allows us to record the fluorescence signal while scanning the electron beam through the optical field of view. By correlating changes in the fluorescence decay with the instantaneous electron beam position and the other EM signals, we obtain a SR fluorescence image (Fig.1). This SR fluorescence image is in perfect registry with the simultaneously acquired EM image.

 

In first experiments on rat pancreas tissue, immuno-labelled for insulin and guanine quadruplexes using different Alexa Fluor dyes, we have achieved a lateral resolution below 100nm (Fig. 2). We will discuss further implementation of our technique towards higher resolution, paving the way towards precise localization, within the EM ultrastructure, of bio-molecules labelled with standard fluorescent dyes.

 

[1] B. Huang, M. Bates, and X. Zhuang, Annual Review of Biochemistry, 78:993-1016, 2009.

[2] P. de Boer, J.P. Hoogenboom, and B.N.G. Giepmans. Nature Methods 12(6):503–513, 2015.

[3] A.C. Zonnevylle et al., Journal of Microscopy 252, 58-70 (2013).

Lennard M. VOORTMAN (Delft, THE NETHERLANDS), Aditi SRINIVASA RAJA, Aaro VÄKEVÄINEN, Pascal DE BOER, Ben N.g. GIEPMANS, Pieter KRUIT, Jacob P. HOOGENBOOM
11:15 - 11:30 #6688 - IM10-OP173 Multi-color correlative PALM/STORM and electron tomography reveals micro-domain organization of endosomes.
Multi-color correlative PALM/STORM and electron tomography reveals micro-domain organization of endosomes.

Correlative Light and Electron Microscopy (CLEM) is a method of choice to demonstrate the localization of a protein in a given organelle while revealing the ultrastructure of the organelle. However, a long-lasting problem in CLEM is the mismatch of resolution between modalities. As it becomes more and more clear that most organelles show a sub-compartmentalization of their membranes into distinct micro-domains, the visualization of such micro-domains at the molecular level using CLEM is a potential strategy to elucidate the molecular basis underlying the ultrastructure of organelles.

We developed a workflow that combines multi-color Single Molecule Localization Microscopy (SMLM) and electron tomography on the very same sample. We applied this approach to study the compartmentalization of early endosomes in micro-domains. As proof of principle, we visualized Transferrin and EGF as models of cargo molecules that follow the recycling and degradative route, respectively. Using single molecule CLEM tomography, we could visualize Transferrin and EGF molecules in distinct morphological compartments within the same endosome (see figure). We could also demonstrate the localization of Rab5 in micro-domains on the globular membrane of the endosome and its exclusion from the tubular parts. Beyond endocytosis, this method can be applied to any biological question requiring both multi-molecule nanoscale localization and 3D ultra-structural information of the very same sub-cellular structure.

Nicolas BROUILLY (Dresden, GERMANY), Yannis KALAIDZIDIS, Jean-Marc VERBAVATZ, Marino ZERIAL
11:30 - 11:45 #5289 - IM10-OP166 Combining SEM with AFM for in situ Correlative Microscopy.
Combining SEM with AFM for in situ Correlative Microscopy.

With the large amount of current research and development focused on nano wires, carbon nano tubes, and other nano scale materials, imaging these materials has become a large part of the challenges involved.

The two most prominent methods for imaging at the nano scale are Scanning Electron Microscopy (SEM) and Atomic Force Microscopy (AFM). These complimentary methods utilize fundamentally different principles for generating imagery - SEM exploits the interaction of electrons with matter, while AFM is based on physical interaction of a sharp tip with the sample surface.

Both approaches have strengths and weaknesses. The SEM's strength is to quickly generate images with a large range of magnifications, making it easy to locate the area of interest. Additionally, the SEM beam can be used to characterize materials beyond mere imaging (e.g. EDX elemental mapping, etc.). Beyond that, the use of Focussed Ion Beams (FIB) enhances the ability for modification or preparation of samples.

However, scanning beams do not yield 3D information, e.g. "invisible" contamination layers or the precise surface structure of novel materials such as solar cells.

The AFM's main advantage lies in its ability to obtain 3D information, the downsides are that it is hard to find the target area and image generation is slow.

Combining these two tools into one setup - putting an AFM inside an SEM - gives quick access to a more complete data set. Additionally, FIB-milled or FIB-deposited structures can be characterized using this combination of tools in a FIB/SEM system.

The utility of this combination of tools is demonstrated with several examples where locating the area of interest purely by AFM or light microscopy would have been highly impractical.

Stephan KLEINDIEK (Reutlingen, GERMANY), Massoud DADRAS, Klaus SCHOCK, Andreas LIEB, Gregor RENKA
11:45 - 12:00 #6983 - IM10-OP176 Probing the multiscale structure, composition and nanomechanical properties of lipids and biopolymers in natural systems: a few examples.
Probing the multiscale structure, composition and nanomechanical properties of lipids and biopolymers in natural systems: a few examples.

The natural systems as plants or animal tissues possess a highly complex organisation of numerous components: micronutriments, proteins, polyssacharides, lipids, water, ….  Their natural arrangement gives very fine structures that range from the nanometer up to few meters scale range (typically from the protein to the whole plant). Native architectures have a direct influence on the functional, chemical, organoleptic and nutritional macroscopic properties relevant for designing sustainable performant products for environmentally compatible food and non–food uses.

The development and application of experimental methodologies to probe the assemblies of lipids, biopolymers, mineral nutrients as well as structural water from the sub-cellular to the molecular levels is then a permanent challenge. The parallel use of a set of complementary microscopy tools that provides relevant information may be needed to overcome the complexity of hydrated natural systems and to get both their structural, composition and nanomechanical properties. Most of the time, the experimental sample preparation is also a critical prerequisite to adapt the biological samples to the features of the analytical tool while keeping as most as possible the structures in their native shape. Thanks to this step, the microscopy tools can be operated through a direct or indirect correlation approach.

Using few applications we developed on biopolymer-based nanoparticles as well as animal cells or plant tissues (see Figure 1), we will show how to probe the structure/composition/nanomechanical properties focusing on the strong complementarity of AFM imaging and force mapping, Raman mapping, SEM/EDX, and scanning transmission X-ray spectrometry (STXM). We will also discuss about the samples preparation strategies, and about data processing in relation to the information expected.

 

 

References

M. Gayral, C. Gaillard, B. Bakan, M. Dalgalarrondo, K. Elmorjani, C. Delluc, S. Brunet, L. Linossier, M.H. Morel, D. Marion, Transition from vitreous to floury endosperm in maize (Zea mays L.) kernels is related to protein and starch gradients, Journal of Cereal Science, 2016, accepté

G. Philippe, C. Gaillard, J. Petit, N. Geneix, M. Dalgalarrondo, R. Franke, C. Rothan, L. Schreiber, D. Marion, B. Bakan, Ester-crosslink Profiling of the Cutin Polymer of Wild Type and Cutin Synthase Tomato (Solanum lycopersicum L.) Mutants Highlights Different Mechanisms of Polymerization, Plant Physiology

C. Karunakaran, C.R. Christensen, C. Gaillard, R. Lahlali, L.M. Blair, V. Perumal, S.S Miller, A.P. HitchcockIntroduction of soft X-ray spectromicroscopy as an advanced technique for plant biopolymers research. PLoS One, 2015 26;10 (3) pages: e0122959.

Covis R., Vives T., Gaillard C., Maud Benoit, Benvegnu T., Interactions and hybrid complex formation of anionic algal polysaccharides with a green cationic glycine betaine derived surfactant, Accepted in Carbohydrate Polymers, 2015, May 5;121:436-48.

Cédric GAILLARD (NANTES)
12:00 - 12:15 #6429 - IM10-OP169 New insights into the Precambrian fossil record using correlative electron and ion beam microscopy.
New insights into the Precambrian fossil record using correlative electron and ion beam microscopy.

Earth’s rock record holds great potential for decoding the origin and early diversification of life on our planet. However, the interpretation of the Precambrian (older than ~541 million years ago) fossil record is fraught with difficulties. These include: the fragmentary nature of the sedimentary rock record with large periods of time unrepresented and certain habitats under-represented; and the nature of the organisms, being microscopic, morphologically simple and often only subtly different from co-occurring non-biological organic material.

Distinguishing between true signs of life and abiotic artefacts requires analytical techniques with excellent spatial resolution in two and three dimensions, in order to accurately analyse key features of putative cells such as cell wall ultrastructure, biochemistry, and interaction of cell walls with the minerals that have fossilised them [1]. Likewise, distinguishing different grades of life (for example, simple prokaryotes versus more complex eukaryotes) requires similar techniques, in order to identify putative multi-cellularity and specific types of cell contents and cell wall architecture. 

We here demonstrate how a protocol combining focused ion beam (FIB) milling, SEM, TEM and nano-scale secondary ion mass spectrometry (SIMS) can reveal unprecedented  nanometer to micrometer scale details of Precambrian fossilised organisms, providing more robust biosignatures for both prokaryotes and eukaryotes for future studies on Earth or other planets. 

FIB milling was used to prepare ultrathin (c. 100 nm) wafers from standard geological thin sections for TEM analysis, plus slightly thicker wafers (c. 150-200 nm) that could be used for both TEM and NanoSIMS analysis. The latter meant that both TEM and NanoSIMS data could be collected from a single candidate microfossil: TEM data included ChemiSTEM elemental mapping of major elements, STEM-EELS analysis of the bonding and structure of organic material, and electron diffraction to identify mineral phases; NanoSIMS data included targeted analysis of trace elements in organic material (e.g., N, S, P) and in the fossilising mineral phases. Analysis of FIB-milled wafers counteracts the problems previously associated with surface analysis techniques such as NanoSIMS (i.e. surface contamination and polishing effects). FIB-milling was also combined with SEM imaging (3D slice and view) in order to obtain accurate 3D visualisations of candidate microfossils.

Data will be presented from three geological formations that play an important role in our understanding of the origin and evolution of early life on Earth: 1, The 1878 Ma Gunflint Formation of Canada, containing an iconic suite of diverse microfossils used as a benchmark for high quality preservation of early life in marine environments [2]; 2, The 1000 Ma Torridon Group of northwest Scotland (Fig. 1) that is renowned for exceptional three-dimensional preservation of both prokaryotes and eukaryotes in phosphate and clay minerals in a terrestrial (lake) setting [3]; 3, The 850 Ma Bitter Springs Formation of central Australia that shows exquisite microfossil preservation (including putative cell contents) in micro-quartz [4].

References cited

[1] D. Wacey et al. (2011) Nature Geosci. 4, 698-702.

[2] D. Wacey et al. (2013) PNAS 110, 8020-8024.

[3] P. Strother et al. (2011) Nature 473, 505-509.

[4] J.W. Schopf (1968) J. Palaeontology 42, 651-688.

Acknowledgements

DW acknowledges funding from the Australian Research Council and European Commission. KE is supported by an Australian Postgraduate Award and a UWA Top-up Scholarship. We acknowledge the facilities, scientific and technical assistance of the AMMRF at UWA, a facility funded by the University, State and Commonwealth Governments. Paul Strother is thanked for provision of the Torridon specimens.

David WACEY, Kate EILOART, Martin SAUNDERS (Perth, AUSTRALIA), Paul GUAGLIARDO, Matt KILBURN
12:15 - 12:30 #6520 - IM10-OP170 Platinum shadowing for correlative light and electron microscopy.
Platinum shadowing for correlative light and electron microscopy.

Fluorescence microscopy reveals molecular expression at nanometer resolution but lacks ultrastructural context information. Electron microscopy provides this contextual subcellular details but protein identification requires elaborate protocols. Correlative light and electron microscopy produces complimentary information that expands our knowledge of protein expression in cells and tissue. Even though a number of correlative approaches are currently available, few of these allow subcellular localization in tissue because of the challenges with sample preparation and 3D complexity. Tokuyasu cryo-sections (Tokuyasu, 1980) preserve the sample ultrastructure and antigenicity of most epitopes, however, heavy metal exposure generates weak contrast to the samples rendering often interpretation of the data difficult.

We present a quick, simple and reproducible method for protein localization by conventional and super-resolution light microscopy combined with platinum shadowing and scanning electron microscopy to obtain topographic contrast from the surface of ultrathin cryo-sections collected on silicon wafers. Figure 1 shows protein distribution at nuclear pores in the topographical landscape of mouse kidney tissue.

Reference: Tokuyasu, K.T. Immunochemistry on ultrathin frozen sections. Histocem. J. 12, 381-403 (1980).

Jose Maria MATEOS (Zurich, SWITZERLAND), Bruno GUHL, Jana DOEHNER, Gery BARMETTLER, Andres KAECH, Urs ZIEGLER

10:15-12:30
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LS5-I
LS5: Extra-cellular matrix
SLOT I

LS5: Extra-cellular matrix
SLOT I

Chairmen: Herman HÖFTE (VERSAILLES, FRANCE), Clemens M. FRANZ (Karlsruhe, GERMANY)
10:15 - 10:45 #8402 - LS05-S16 Structure and nanomechanical properties of a wonderfully complex material, the primary cell wall of plants: recent progress based on AFM and FESEM.
Structure and nanomechanical properties of a wonderfully complex material, the primary cell wall of plants: recent progress based on AFM and FESEM.

Growing plant cells synthesize a strong yet extensible cell wall (=extracellular matrix) composed of long, thin cellulose microfibrils that are laterally bonded to one another in organized layers ~ 40 nm thick and that are embedded in a hydrated matrix consisting of complex polysaccharides (pectins, hemicelluloses). I will briefly review the biosynthetic origins of these wall components and what is known about their assembly to form a hierarchically-structured hydrated material with diverse physical and chemical properties. Recent advances in atomic force microscopy (AFM), field effect scanning electron microscopy (FESEM), and solid-state NMR have led to a rethinking of how these wall components interact with one another and how cells regulate the irreversible expansion (growth) of the cell wall 1-4. With AFM we have characterized the detailed organization of recently-deposited cellulose microfibrils in never dried cell walls of onion epidermis. The interaction of microfibrils with matrix can be visualized in two-color maps based height (which emphasizes microfibrils and modulus or adhesion, which highlight soft matrix. Additionally, when the cell walls are stretched to reveal microfibril re-arrangements after plastic or elastic deformation or after enzyme-mediated cell wall creep, we find different patterns of microfibril movements. FESEM images of the same material (except dried) detect only a surface layer of pectins that obscure the underlying microfibrils. These can be unveiled by enzyme digestions to selectively remove pectins. Some of the remaining unsolved problems in cell wall structure and polysaccharide interactions will be highlighted.

 References:

1 Zhang T, Zheng Y, Cosgrove DJ. (2016) Spatial organization of cellulose microfibrils and matrix polysaccharides in primary plant cell walls as imaged by multichannel atomic force microscopy. Plant J  85: 179-92.

2  Wang T, Park YB, Cosgrove DJ, Hong M. (2015) Cellulose-pectin spatial contacts are inherent to never-dried arabidopsis primary cell walls: Evidence from solid-state nuclear magnetic resonance. Plant Physiol  168: 871-84.

3 Cosgrove DJ. (2015) Plant cell wall extensibility: Connecting plant cell growth with cell wall structure, mechanics, and the action of wall-modifying enzymes. J Exp Bot  67: 463-476.

4 Cosgrove DJ. (2014) Re-constructing our models of cellulose and primary cell wall assembly. Curr Opin Plant Biol  22C: 122-31.

 Acknowledgements: This work was supported by the Center for Lignocellulose Structure and Formation, an Energy Frontier Research Center funded by the U.S. Department of Energy, Office of Science, Basic Energy Sciences (grant no. DE-SC0001090).

Daniel COSGROVE (, USA), Tian ZHANG, Yunzhen ZHENG
Invited
10:45 - 11:15 #8783 - LS05-017 Mechanical Properties of the Extracellular Matrix Affect Growth Plate Morphogenesis.
Mechanical Properties of the Extracellular Matrix Affect Growth Plate Morphogenesis.

The growth plate (GP) is a dynamic tissue driving bone elongation through chondrocyte proliferation, hypertrophy and matrix production. The extracellular matrix (ECM) is the major determinant of GP biomechanical properties and assumed to play a crucial role for chondrocyte geometry and arrangement, thereby guiding proper growth plate morphogenesis and bone elongation. To elucidate the relationship between morphology and biomechanics during cartilage morphogenesis, we have investigated structural and elastic properties of the proliferative zone of the murine GP by atomic force microscopy (AFM) from the embryonic stage to adulthood. We observed a progressive cell flattening and arrangement into columns from embryonic day 13.5 until postnatal week 2, correlating with an increasing collagen density and ECM stiffness, followed by a nearly constant cell shape, collagen density and ECM stiffness from week 2 to 4 months. At all ages, we found marked differences in the density and organization of the collagen network between the intracolumnar matrix, and the intercolumnar matrix, associated with a roughly two-fold higher stiffness of the intracolumnar matrix compared to the intercolumnar matrix. This difference in local ECM stiffness seems therefore to force the cells to arrange in a columnar structure upon cell division and drive bone elongation during embryonic and juvenile development.

References:

C. Prein, N. Warmbold, Z. Farkas, M. Schieker, A. Aszodi, and H Clausen-Schaumann: Structural and Mechanical Properties of the Proliferative Zone of the Developing Murine Growth Plate Cartilage Assessed by Atomic Force Microscopy, Matrix Biology, (2016), 50, 1-15.

M. Kamper, N. Hamann, C. Prein, H Clausen-Schaumann, Z. Farkas, A. Aszodi, A. Niehoff, M. Paulsson, and F. Zaucke: Early changes in morphology, bone mineral density and matrix composition of vertebrae lead to disc degeneration in aged collagen IX -/- mice, Matrix Biology, (2016), 49, 132-143.

E. Aro, A. M. Salo, R. Khatri, M. Finnilä, I. Miinalainen, R. Sormunen, O. Pakkanen, T. Holster, R. Soininen, C. Prein, H. Clausen-Schaumann, A. Asźodi, J. Tuukkanen, K. I. Kivirikko, E. Schipani, and J. Myllyharju: Severe Extracellular Matrix Abnormalities and Chondrodysplasia in Mice Lacking Collagen Prolyl 4-Hydroxylase Isoenzyme II in Combination with a Reduced Amount of Isoenzyme I,  J. Biol. Chem., (2015), 290 (27), 16964–16978.

Carina PREIN, Niklas WARMBOLD, Zsuzsanna FARKAS, Matthias SCHIEKER, Attila ASZODI, Hauke CLAUSEN-SCHAUMANN (, GERMANY)
11:15 - 11:45 #8298 - LS05-S18 Second harmonic imaging of collagen organization in connective tissues.
Second harmonic imaging of collagen organization in connective tissues.

Type I collagen is a major structural protein in mammals. This biopolymer is synthesized as a triple helix, which self-assembles into fibrils (diameter: 10-300 nm) and further forms various 3D organizations specific to each tissue. In recent years Second Harmonic Generation (SHG) microscopy has emerged as a powerful technique for the in situ investigation of the fibrillar collagen structures in matrices or tissues [1]. However, as an optical technique with typically 300 nm lateral resolution, SHG microscopy cannot resolve most of the collagen fibrils. Moreover, in contrast to incoherent fluorescence signals that scale linearly with the chromophore concentration, SHG is a coherent multiphoton signal that scales quadratically with the density of collagen triple helices aligned with the same polarity in the focal volume. Consequently, quantitative SHG measurements have been limited so far to averaged phenomenological parameters [1].

In this study, we correlated SHG and transmission electron microscopies to determine the sensitivity of SHG microscopy and calibrate SHG signals as a function of the diameter of the collagen fibril [2]. To that end, we synthesized in vitro isolated fibrils with various diameters and successfully imaged the very same fibrils with both techniques, down to 30 nm diameter (see figure 1). We observed that SHG signals scale as the fourth power of the fibril diameter, as expected from analytical and numerical calculations. It validated our quantitative bottom-up approach used to calculate the non-linear response at the fibrillar scale and demonstrated that the high sensitivity of SHG microscopy originates from the parallel alignment of triple helices within the fibrils and the subsequent constructive interference of SHG radiations. This calibration was then applied to intact rat corneas, where we successfully recovered the diameter of hyperglycemia-induced fibrils in the Descemet’s membrane without having to resolve them [2,3].          

Importantly, this calibration only applies to isolated fibrils. Nevertheless, complementary techniques can probe the sub-micrometer structure of dense distributions of collagen fibrils. In particular, we have shown that polarization-resolved SHG microscopy can probe the main orientation of collagen fibrils and their orientation disorder within the focal volume [4]. Combination of this modality with traction assays then provides a new method to measure the reorganization of the collagen network upon stretching and to correlate this microscopic response to the biomechanical response at macroscopic scale [4, 5].

In conclusion, our data represent a major step towards quantitative SHG imaging of collagen organization in biomaterials or connective tissues.

 

[1] M. Strupler, A.-M. Pena, M. Hernest, P.-L. Tharaux, J.-L. Martin, E. Beaurepaire, and M.-C. Schanne-Klein, Opt. Express 15, 4054-4065 (2007).

[2] S. Bancelin, C. Aimé, I. Gusachenko, L. Kowalczuk, G. Latour, T. Coradin, and M.-C. Schanne-Klein, Nat. Commun. 5 (2014).

[3] G. Latour, L. Kowalczuk, M. Savoldelli, J.-L. Bourges, K. Plamann, F. Behar-Cohen, and M.-C. Schanne-Klein, PLos ONE 7, e48388 (2012).

[4] I. Gusachenko, Y. Goulam Houssen, V. Tran, J.-M. Allain, and M.-C. Schanne-Klein, Biophys. J. 102, 2220 (2012).

[5] S. Bancelin, B. Lynch, C. Bonod-Bidaud, G. Ducourthial, S. Psilodimitrakopoulos, P. Dokladal, J.-M. Allain, M.-C. Schanne-Klein, and F. Ruggiero, Scientific Reports 5, 17635 (2015)

Marie-Claire SCHANNE-KLEIN (PALAISEAU CEDEX)
Invited
11:45 - 12:00 #6001 - LS05-OP023 Interplay of organic matrix and amorphous calcium phosphate strengthens the isopod claw.
Interplay of organic matrix and amorphous calcium phosphate strengthens the isopod claw.

Animal skeletons are high-performing composite materials that may help inspire materials and designs in a broad spectrum of industrial and biomedical applications. The study of various skeletal elements can provide important insights into evolutionary solutions to mechanical demands of animal locomotion, feeding and reproduction, as well as reveal the mechanisms controlling skeletal formation and biomineralization.

The extracellular matrix forming the crustacean exoskeleton comprises chitin-protein fibers embedded in a calcified inorganic matrix that consists of calcite and amorphous calcium carbonate. In crustaceans, the cuticle can be subdivided into a thin external layer - the epicuticle – and two internal layers, which are heavily calcified – the exocuticle and the endocuticle [1]. The crustacean cuticle generally consists of stacked sheets of parallel chitin-protein fibers, which helicoidally shift their orientation in each sequential sheet, resulting in a structure referred to as the Bouligand pattern [2]. This organization of the fibers strengthens the cuticle in different directions.

 

In our study, we analyzed the structure and composition of the walking leg claw of the woodlouse Porcellio scaber. Woodlice are terrestrial crustaceans that support their bodies with 7 pairs of legs, each ending in a claw. The claws are thin skeletal elements predominantly subjected to unidirectional loads. To study the nano-structure of the matrix, we imaged fractured claws with field-emission scanning electron microscopy using a Jeol 7500F microscope. We then analyzed the elemental composition and the distribution of mineral components in the claws with energy-dispersive X-ray spectroscopy (EDX) and electron energy-loss spectroscopy (EELS) combined with high-angle annular dark field scanning transmission electron microscopy (HAADF-STEM) at high spatial and high energy resolution using Zeiss SESAM and Jeol ARM200F microscopes at different accelerating voltages.

 

Our results demonstrate that the exocuticle of the claw is not calcified and is heavily brominated instead. The endocuticle, on the other hand, is mineralized predominantly with stable amorphous calcium phosphate, which is a highly unusual feature of an animal exoskeleton. Furthermore, we established that the claw endocuticle is highly structurally anisotropic, consisting of axially oriented chitin-protein fibers and amorphous calcium phosphate particles, all oriented in the direction of loading.

 

The presence of amorphous calcium phosphate in the mineralized endocuticle and a non-calcified, brominated external exocuticle may help increase fracture resistance of the claw cuticle. The brominated exocuticle, which is likely more elastic than the mineralized endocuticle, is distributed in areas subjected to maximum stress during axial loading of the claw. These structural and compositional features of the claw cuticle likely result in greater resistance of the claw to fracture and wear when exposed to axial loading.

 

Acknowledgments

The research leading to these results has received funding from the European Union Seventh Framework Program [FP/2007-2013] under grant agreement No.312483 (ESTEEM2). The work was supported by the Slovenian Research Agency in the scope of the research program P1-0184 (Integrative zoology and speleobiology).

 

References

[1] R Roer and R Dillaman, American Zoologist 24 (1984), pp 893-909.

[2] Y Bouligand, Tissue & Cell 4 (1972), pp. 189-217.

Miloš VITTORI (Ljubljana, SLOVENIA), Vesna SROT, Birgit BUSSMANN, Peter A. VAN AKEN, Jasna ŠTRUS
12:00 - 12:15 #6349 - LS05-OP024 Enamel evolution: Back in time by a molecular manipulation.
Enamel evolution: Back in time by a molecular manipulation.

Biomineralization is one of the key processes during vertebrate evolution that incorporates calcium and phosphate ions into soft matrices in the form of hydroxyapatite. Occurrence of mineralized tissues have offered the basis for various adaptive phenotypes such as endoskeleton for locomotion (bone), body armor for protection and teeth (enamel, dentin) for predation. Enamel is unique among the mineralized tissues, as it is formed on specific network of enamel matrix proteins (EMPs) secreted by epithelial ameloblasts. The two key structural proteins of enamel, amelogenin (AMEL) and ameloblastin (AMBN), self-assemble into higher-ordered structures from monomeric intrinsically disordered subunits as does the type I collagen (COL1), the predominant matrix protein in bone and dentine. While COL1 undergoes self-assembly via consecutive Gly-X-Y motif, the mechanism of self-assembly of EMPs and their subsequent role in formation of organized layer of hydroxyapatite crystals remains poorly understood.

We report here a novel evolutionary conserved self-assembly motif common to the key structural enamel matrix proteins AMBN and AMEL. The presence of this motif is essential for self-assembly of AMBN and AMEL into higher-ordered structures. These structures are essential for proper enamel formation. Transgenic mice that were unable to produce supramolecular structures of AMBN due to point substitutions within the identified self-assembly motif then produced severely affected enamel with simplified organization. Despite the normal cellular organization, EMPs secretion and Ca and P levels within the growing enamel of transgenic mouse, the affected enamel lacked organized prismatic structures and showed only radial organization without visible Hunter-Schreger bands. Moreover, enamel of mutant mice contained an enormous portion of interprismatic matrix with hypomineralized, yet well recognizable, crystallites, while no formation of oriented crystallites was observed within the compromised prisms.

This is the first in vivo evidence that the formation of supramolecular structures of enamel matrix proteins was essential for evolution of highly structured enamel in mammals.

Tomas WALD (Prague 10, CZECH REPUBLIC), Frantisek SPOUTIL, Adriana OSICKOVA, Michaela PROCHAZKOVA, Oldrich BENADA, Petr KASPAREK, Ladislav BUMBA , Ophir KLEIN, Radislav SEDLACEK , Peter SEBO, Jan PROCHAZKA, Radim OSICKA
12:15 - 12:30 #6907 - LS05-OP025 The sample preparation for cryo-SEM: the real ultrastructure of microbial biofilm or just artifacts?
The sample preparation for cryo-SEM: the real ultrastructure of microbial biofilm or just artifacts?

The cryo-scanning electron microscopy (cryo-SEM) belongs to reputable techniques in electron microscopy of hydrated samples such as biofilms. The crucial steps of the cryo-preparation techniques are primarily the cryo-fixation and partial sublimation of ice contamination caused during the transfer of the sample to the cryo-high-vacuum preparation chamber where the sublimation process is performed; optionally the freeze-fracturing or coating by metal sputtering or carbon evaporation can be applied. In the case of cryo-fixation, an effort is to keep the frozen biofilm in the form nearby its native state. One of the simplest cryo-fixation techniques is a plunging of the biofilm on a substrate into a liquid cryogen. However, the plunging into a liquid nitrogen or even liquid ethane/propane is sufficient for fixation of very thin layers of biofilm (no more than a few micrometers in thickness) because it is very difficult to achieve enough cooling rates to produce amorphous ice in the sample due to the Leidenfrost effect [1]. Moreover, we show that the cryo-fixation into liquid nitrogen can lead to significant lateral macro-segregation of both bacteria and extracellular polymeric substances (EPS), where plunging into liquid ethane leads to micro-segregation of EPS and macro-segregation of bacteria (Figure 1, 2A). Substantially more effective cooling can be achieved by increasing the pressure during exposure to the liquid cryogen. This can be performed for example by the high-pressure freezing (HPF) technique [2]. It was proved that cryo-fixed biofilms by HPF show significantly improved preservation of bacterial ultrastructure and biofilm organization (Figure 2B).

In this study, the multi-layered biofilms formed by microorganisms were observed by cryo-SEM using freeze-fracturing technique. Cryo-fixation methods like plunging into liquid cryogen, freezing by cryo-jet system and high pressure freezing are compared. The freeze-fracture technique consists of fracturing a rapidly frozen biological sample; structural details exposed by the fracture plane may be then visualized by cryo-SEM.

The well-characterized ica operon-positive, biofilm and slime producing Staphylococcus epidermidis strain CCM 7221 (Czech Collection of Microorganisms, Brno, Czech Republic)  and Candida parapsilosis BC11 from Collection of Microbiology Institute, Masaryk University and St. Anna University Hospital (Brno, Czech Republic) was observed.  The strains Candida albicans GDH 2346 were also included [3]. Cultures were cultivated on the sapphire discs or cover glass in the cultivation BHI medium at 37°C for two days; fractured after cryo-fixation, then followed short sublimation of ice contamination at -90°C which, moreover, partially exposes interior of the biofilm. In our experiments we focused on the formation of the extracellular matrix produced during the cultivation.

The cryo-fixation can be recognized as a sufficient way how to fix and stabilize biofilms before their examination in cryo-SEM. The simple plunging into liquid cryogens is applicable only for very thin specimens depending on the composition and used substrate. In this case of grown biofilms which thickness is usually more than 10 µm then the inner structure of matrix and bacteria interconnections were observed applying the freeze-fracture technique was used, the HPF technique has proved to be necessary for preserving the biofilm ultrastructure.

References:

1.             Kuo, J., Electron microscopy: Methods and protocols. Vol. 369. 2007: Springer Science & Business Media.

2.             Krzyzanek, V., et al., Cryo-SEM of perpendicular cross freeze-fractures through a high-pressure-frozen biofilm. Microscopy and Microanalysis, 2014. 20(3): p. 1232-1233.

3.             Ruzicka, F., et al., Importance of biofilm in Candida parapsilosis and evaluation of its susceptibility to antifungal agents by colorimetric method. Folia Microbiol (Praha), 2007. 52(3): p. 209-14.

This work received support from the Grant Agency of the Czech Republic (GA14-20012S and GA16-12477S). KH acknowledges the support of FEI/CSMS scholarship.

Kamila HRUBANOVA (Brno, CZECH REPUBLIC), Radim SKOUPY, Jana NEBESAROVA, Filip RUZICKA, Vladislav KRZYZANEK

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AWARDS
EMS AWARDS

EMS AWARDS

14:00 - 15:30 Exit Wavefunction Reconstruction:  Current Status, Future Prospects and Applications to Materials Systems. Angus KIRKLAND (Oxford, UK)
14:00 - 15:30 Scanning Transmission Electron Microscopy of Eukaryotic Cells in Liquid. Niels DE JONGE (Saarbrücken, GERMANY)
14:00 - 15:30
These lectures will be followed by the Outstanding Paper Award ceremony.