Mikroskopie 2011 Československá mikroskopická společnost Hotel SKI, Nové Město na Moravě, února 2011

Mikroskopie 2011

Československá mikroskopická společnost Hotel SKI, Nové Město na Moravě, 17. – 18. února 2011

Synura petersenii, planktonní řasa, patří k autotrofním zástupcům chromist. Jemné struktury křemičitých šupin, které kryjí povrch organismu, jsou důležitými taxonomickými znaky pro vnitrodruhovou klasifikaci. Vzorek přilepený na polykarbonátovém filtru poly-L-lyzinem byl vysušen kritickým bodem, pokoven zlatem ve sputter-coateru a zobrazen v elektronovém mikroskopu Philips CM12/STEM v rastrovacím módu pomocí detektoru sekundárních elektronů při urychlovacím napětí 80 kV. Snímek vznikl ve spolupráci s Katedrou botaniky, PřF UK v Praze. Autor snímku: Oldřich Benada, MBÚ AV ČR, v.v.i.

2

Mikroskopie 2011 Pořádá: Československá mikroskopická společnost Vídeňská 1083, 142 20 Praha 4 Tel./Fax +420-241 062 219 email: [email protected] www: http:/www.microscopy.cz

Programoví organizátoři: RNDr. Luděk Frank (fyzika, materiály, přístroje...) email: [email protected] Prof. Pavel Hozák (biologie, medicína,...) email: [email protected]

Hotel SKI, Nové Město na Moravě 17. – 18. února 2011 3

Sponzorují:

4

Program Čtvrtek 17. února 10:00 - 11:30 registrace 11:30 - 12:30 oběd 12:30 - 12:40 zahájení – Pavel Hozák, předseda ČSMS 12:40 - 12:50 vyhlášení ceny ČSMS pro rok 2010 za zásluhy v mikroskopii: cenu získal František Lednický za celoživotní působení v oboru elektronové mikroskopie materiálů 12:50 - 13:20 přednáška laureáta: František Lednický: Mikrosvět polymerních materiálů [1] 13:20 - 13:25 Vyhlášení vítěze soutěže za nejlepší obrázky pro www IMC 2014 13:25 - 13:30 Vyhlášení soutěže o stipendium FEI - CSMS I. blok přednášek – zvané přednášky (moderátor: Pavel Hozák) 13:30 - 14:00 Petr Šittner: Nonconventional thermomechanical processing of thin NiTi filaments by electric current – TEM investigation of defects and microstructures [2] 14:00 - 14:30 Martin Anger: Using live cell imaging to study molecular mechanisms of chromosome segregation in meiosis [3] 14:30 - 15:00 Matej Pospiech: Mikroskopia potravín [4] 15:00 - 16:30 postery a firemní výstava s občerstvením 16:30 - 18:00 valné shromáždění ČSMS 18:30 - 19:30 řízená ochutnávka vína z produkce vinařství Lahofer www.vinarstvi-lahofer.cz 19:30 - 23:00 společenský večer s rautem a hudbou skupiny Lucie Revival www.lucierevival.com

5

Pátek 18. února II. blok přednášek - materiálové vědy (moderátor: Ivo Vávra) 9:00 - 9:15 Mariana Klementová: Structure of η’-Phase of Cu3+x(Si,Ge) determined by quantitative electron diffraction tomography [5] 9:15 - 9:30 Peter Švec: Interface evolution study in rapidly quenched amorphous bilayers [6] 9:30 - 9:45 Václav Tyrpekl: Microscopy analyses for the behavior description of melt systems formed during nuclear reactor severe accident [7] 9:45 - 10:00 Niva Zárubová: Mechanism of twinning in NiMnGa shape memory alloy studied by IN-SITU TEM straining [8] III. blok přednášek - mikroskopická instrumentace (moderátor: Luděk Frank) 10:00 - 10:15 Regina Holčáková: Innovative research in electron microscopes, analysis of magnetic field distribution of some types of magnetic lenses by FEM [9] 10:15 - 10:30 Miroslav Kolíbal: Watching nanowires grow [10] 10:30 - 10:45 Jaroslav Lukeš: In-situ SEM and TEM nanomechanical testing of materials [11] 10:45 - 11:00 Ilona Müllerová: Very low energy STEM [12] 11:00 - 11:30 přestávka IV. blok přednášek – firemní prezentace nových přístrojů/technik (moderátor: Fedor Čiampor) 11:30 - 11:45 CARL ZEISS - Pavel Krist: Spectral Imaging and Linear Unmixing [13] 11:45 - 12:00 CARL ZEISS - Peter Gnauck: High resolution large area Imaging of biological samples and correlative microscopy [14] 12:00 - 12:15 FEI - Tomáš Vystavěl: Novinky z FEI 12:15 - 12:30 MIKRO – Zdeněk Rous: Konfokální mikroskopie LEICA – Novinky 2010 [15] 12:30 - 13:30 oběd 13:30 - 13:45 NIKON - Ivan Rozkošný: Confocal systems and super resolution Nikon [16] 6

13:45 - 14:00 PE SYSTEMS - Radovan Horák: PerkinElmer – Live cell imaging data management [17] 14:00 - 14:15 SPECION - Mark Richter: The Nanowizard 3 – The most flexible, high resolution AFM with true optical integration [18] 14:15 - 14:30 TESCAN - Kristýna Kubíčková: Biological applications of the scanning electron microscope MIRA 3 FEG SEM [19] V. blok přednášek - biologie a medicína (moderátor: Lucie Kubínová) 14:30 - 14:45 Jan Klepetář: Confocal fluorimetry – analytical approach [20] 14:45 - 15:00 Oldřich Benada: Analysis of nanoparticles in soft oral tissues [21] 15:00 - 15:15 Radek Pelc: Artifact-reduction strategies in biological phase-contrast microscopy [22] 15:15 - 15:30 Margarita Sobol: How to improve preparation of samples for immunogold labeling on ultrathin sections? [23] 15:30 - 15:45 Pavel Hozák: Multiple immunolabeling of antigens on resin sections of biological specimens [24] 15:45 - 16:00 zakončení

7

PŘEDNÁŠKY (uspořádáno podle programu)

8

1 MICROWORLD OF POLYMERIC MATERIALS Lednický F.1,2 1 2

Institute of Macromolecular Chemistry, AS CR, Prague; Technical University of Liberec, Faculty of Mechanical Engineering.

Annual worldwide volume production of polymeric materials exceeds substantially the contemporary production of metallic materials. However, no adequate attention of scientists and technicians is devoted to polymers. Long macromolecular chains of polymers form characteristic structures, which are the grounds of the specific macroscopic properties. Crystalline textures are prevailingly formed from folded chains crystals separated by the amorphous phase of disordered macromolecular chains or their parts. Higher-order formations typical for solid polymeric crystalline materials are spherulite, which arise during the cooling of melt, the most frequent procedure in fabrication of final polymer products. Combining polymeric materials into blends, and also filling or reinforcing the polymer matrix with non-polymeric particulate materials make it possible to produce tailored materials with requested properties. The most important ways to study the supermolecular structure (morphology) of polymers are every microscopy techniques. Variability of polymer properties (from glassy-hard to rubber-like soft) requests the application of the microscopy preparation techniques, which are used in all the scientific disciplines, from the techniques specific for ceramic or metallic materials to those used in biology or medicine. In addition to the basic knowledge of the phase arrangements (crystalline, amorphous, distribution of the individual polymeric constituents, distribution of filler or the reinforcement phase), microscopic techniques are a powerful tool to reveal processing defects, flaws, or the failure sources. That knowledge then contributes greatly to formulate new polymeric materials with the required properties, which can be reached owing to the proper chemical and supermolecular structure.

9

2 NONCONVENTIONAL THERMOMECHANICAL PROCESSING OF THIN NITI FILAMENTS BY ELECTRIC CURRENT – TEM INVESTIGATION OF DEFECTS AND MICROSTRUCTURES Šittner P.1 , Dellvile R.2, PIlch J.1 1

Institute of Physics of the ASCR, Czech Republic; 2EMAT, University of Antwerp, Belgium. Thin filaments made of NiTi shape memory alloy are currently used in smart technical textiles, developed especially for applications in medicine. These metallic filaments exhibit unique physical and mechanical properties which are not commonly encountered in conventional textile materials as high strength, nonlinear superelastic stress-strain behavior, shape memory effect, electrical conductivity, unique response to external stimuli as thermal actuation or impact loads etc. Recently, we have applied a nonconventional heat treatment by electric current (FTMT-EC method [1,2]) to prepare thin NiTi filaments (d=20-100μm) with unique nanosized microstructure (grain size 20200nm) and superior functional properties. Obviously, the relationship between the FTMT-EC process parameters, microstructure and functional properties are of key importance for development of filament processing technology. In this work we will discuss the results of systematic TEM [3,4] and synchrotron X-ray [2] investigations of microstructures and defects in heat treated [3] and mechanically cycled [4] NiTi filaments. References [1] J.Pilch, L.Heller and P.Sittner ASM International, SMST E-elastic Newsletter, January 2010 [2] B. Malard, J. Pilch, P. Sittner, R. Delville, C. Curfs, Acta materialia, 59, 2011, 1542–1556 [3] R. Delville, B. Malard b, J. Pilch, P. Sittner, D. Schryvers Acta Materialia 58 (2010) 4503–4515 R. Delville, B. Malard, J. Pilch, P. Sittner, D. Schryvers, International Journal of Plasticity 27 (2011) 282–297

10

3 USING LIVE CELL IMAGING TO STUDY MOLECULAR MECHANISMS OF CHROMOSOME SEGREGATION IN MEIOSIS ANGER M.1,2 1

Institute of Animal Physiology and Genetics, Rumburska 89, Libechov 277 21, Czech Republic 2 Veterinary Research Institute, Hudcova 70, Brno 621 00, Czech Republic Mammalian oocytes are unique cells, which first appear during early embryonic development. After the initial stages of meiotic program, involving also recombination and exchange of genetic material between homologous chromosomes, oocytes are arrested in prophase of the first meiotic division until hormonal stimulation triggers resumption of meiosis. The list of techniques, which could be used to study chromosome segregation during meiosis in mammalian oocytes, is rather limited. The main reason is that the number of cells, which can be obtained, is usually very low, ruling out biochemical techniques for analysis of their protein content. On the other hand, oocytes are large and fairly transparent, which is why they represent ideal objects for live cell imaging. In a relatively simple culture system that can be easily reproduced on the microscope they are also able to complete meiosis in vitro. By employing ZP3-Cre lox system for gene targeting, or microinjection of RNA or proteins for overexpression, we can achieve changes in the expression level of a particular gene specifically in oocytes without interfering with other cells in the organism. All these techniques together can be used to study not only chromosome segregation during both meiotic divisions, but also changes of the activity of enzymatic complexes in living cells.

11

4 FOOD MICROSCOPY Pospiech M., Tremlová B., Talandová M., Randulová Z., Řezáčová Lukášková Z. Department of Vegetable Foodstuffs and Plant Production, VFU Brno. Microscopic methods have a long tradition in the analysis of foodstuffs. The first used method was detection of adulteration in coffee. In present day, microscopy methods are mainly used for determination of composite food ingredients, determination of the quality and arrangement of food components, detection of foodstuffs adulteration, detection of risky food components and, in case of using histometry, stereology and image analysis, also for the quantification of food ingredients. Food microscopy applies a rich range of microscopy methods. Light microscopy is used for determination of food ingredients mainly in meat products. Fluorescent methods are used in determination of vegetable products. Some of these microscopy methods use autoflorescence of plant cells. Both of these methods are used for microstructure description. For ultramorphology of foodstuff, TEM and SEM were useful. These methods provide information about relations between proteins, fats and other components that may explain their relationship in food matrices. In appropriate cases, microscopic methods are used as technical studies necessary for food production, but they are also used by supervisory authorities in the field of food safety. The project was supported by MŠMT č. 6215712402 and IGA 86/2010FVHE (for more information see: www.foodmicroscopy.com).

12

5 STRUCTURE of η’-PHASE of Cu3+x(Si,Ge)DETERMINED BY QANTITATIVE QUANTITATIVE ELECTRON DIFFRACTION TOMOGRAPHY Klementová M.1,2, Palatinus L.1, Dřínek V.3 Jarošová M.1, Petříček V.1

1

Institute of Physics of the AS CR, v.v.i., Prague; 2 Institute of inorganic chemistry of the AS CR, v.v.i., Husinec-Řež; 3 Institute of Chemical Process Fundamentals of the AS CR, v.v.i., Prague, Czechia Small isolated platelets about 40 nanometres thick with composition η’Cu3+x(Si,Ge) were obtained by deposition of organometallic precursors on Cu substrate by the CVD method. The platelets were analyzed by quantitative electron diffraction tomography coupled with precession electron diffraction, and the structure was solved by the charge-flipping algorithm in superspace. The structure is trigonal, and it is incommensurately modulated with two modulation vectors q1 = (α,α,1/3) and q2 = (-2α,α,1/3), superspace group P 3 1m (α,α,1/3) 000 (-2α,α,1/3) 000. It is formed by slabs of Cu clusters separated by monoatomic layers of Si and Ge atoms. The Cu slabs are strongly modulated, leading to a predominant icosahedral coordination of the central Cu atoms. The twodimensional modulation functions describing the shifts of the atoms show an unprecedented complexity and large amplitude. The conflict between the strive for locally favorable icosahedral coordination of the Cu atoms and the need for a long-period arrangement is the most likely reason for the modulation.

13

6 INTERFACE EVOLUTION STUDY IN RAPIDLY QUENCHED AMORPHOUS BILAYERS Švec P., Maťko I., Janičkovič D., Švec Sr. P. Institute of Physics, Slovak Academy of Sciences, Bratislava, Slovakia The interface between two amorphous layers with different chemical composition has been studied on rapidly quenched bilayers consisting of FeSi-B and Co-Si-B, respectively. Fully amorphous bilayer samples were prepared by planar flow casting using double-nozzle technique. Temperature dependencies of electrical resistivity, dilatation and magnetization have been investigated in the amorphous state and during crystallization of both layers, which take place at different temperatures. The structure of the interlayer has been investigated by cross-sectional TEM, XRD, SEM/EDX. Originally narrow interface of submicron thickness remains unchanged with increasing annealing temperature all the way up to the crystallization of the more stable layer. Above this temperature interval where both layers have transformed into crystalline state, the interface changes its character and thickness. Forrmation of phases which are structurally and morphologically different from those formed in the bulk of each layer, is observed. This indicates that the atomic diffusion process in amorphous state is significantly slower than that in the corresponding crystalline counterparts. More detailed analysis and quantification of the evolution of elemental distributions in the bilayer interface area provides an important tool for investigations of the differences in mechanisms of mass transport in rapidly quenched amorphous matrix as compared to the classical diffusion mechanisms in polycrystalline alloys. Support of the Agency of the Ministry of Education of the Slovak Republic for the Structural Funds of the EU (CEKOMAT I, ITMS 26240120006) is gratefully acknowledged. The work has been supported in part also by the project VEGA 2/0111/11.

14

7 MICROSCOPY ANALYSES FOR THE BEHAVIOR DESCRIPTION OF MELT SYSTEMS FORMED DURING NUCLEAR REACTOR SEVERE ACCIDENT Tyrpekl V.1,2, Piluso P.2, Bakardjieva S.1,Kiselová M.3 1

Institute of Inorganic Chemistry, v.v.i. Academy of Science of the Czech Republic, Husinec-Řez 1001, Řez 25068, Czech Republic ; 2 CEA, DEN, STRI/LMA, Cadarache, F-13108 St Paul lez Durance; 3 Nuclear Research Institute UJV Ltd., Řez 25068, Czech Republic. For the safety evaluations of recent and future nuclear power-plants basic phase behavior description of the materials are necessary. In the case of nuclear reacor core melt-down accident a complex melt mixture, called corium, is formed. This melt mixture could react and ablate the surrounding material structure endangering the reactor vessel and containment integrity. Interaction of the core melt with water, concrete and steel has been widely studied in the past. This contribution is focused on the work done in the cooperation of UACH AV CR,v.v.i and UJV Řež a.s. in the field of phase behavior of prototypical core melt (UO2-ZrO2 mixture) and iron oxides, concrete and water. Solid state analyses of solidified melts provide basic data for the phase diagram construction and solidification path reconstruction. Analyses of materials simulating the core melt, facilitating the materal handling (like Al2O3, tin, etc.), in the experiments will be also presented. These results help to reveal the physico-chamical phenomena during the processes and can be used in safety codes devoted to the nuclear reactor severe accident.

15

8 MECHANISM OF TWINNING IN NiMnGa SHAPE MEMORY ALLOY STUDIED BY IN-SITU TEM STRAINING Zárubová N. 1, Ge Y.2, Gemperlová J. 1, Gemperle A.1, Dlabáček Z.1, Hannula S.-P.2 1

Institute of Physics, ASCR v.v.i., Prague, Czech Republic Aalto University, Dpt. of Materials Science and Engineering, FI-02150 Espoo, Finland

2

Mobility of twin boundaries plays a highly important role in the thermally induced shape memory effect, superelasticity and magnetic-field-inducedstrain in NiMnGa alloys. In the present study, in-situ straining in a transmission electron microscope was performed to follow the twinning/detwinning processes in the non-modulated tetragonal martensite [1]. The tensile straining tests were carried out in a JEM 1200EX microscope equipped with a double tilt straining stage. The martensitic structure of the samples before straining consisted of thermally induced self-accommodated multi-variants. During the in-situ straining, detwinning processes were observed and video-recorded. The volume fraction of the twin variants which are more favorably oriented to the applied tensile stress increased at the expense of the less favorably oriented ones. Under special conditions, the reverse process (twinning) took place. A detailed structural study of the strained samples was carried out in the JEM and in a Tecnai F20 G2 200kV FEG microscope. The twinning/detwinning processes occur by movement of twinning dislocations emitted from the boundaries between individual twinned regions. The boundaries serve at the same time as sinks for the progressing twinning dislocations. The twin dislocatons glide along the {101} twin planes and their Burgers vector is parallel to the <101> directions. Computer simulations of the dislocation contrast are now being done with the aim to determine the size of the Burgers vector of the twinning dislocations, and to elucidate in detail the mechanism of the twinning/detwinning process. [1] Y. Ge, N. Zárubová, Z. Dlabáček, I. Aaltio, O. Söderberg, S.-P. Hannula, Proceedings of ESOMAT 2009, 04007 (2009) DOI: 10.1051/esomat/200904007.

16

9 INNOVATIVE RESEARCH IN ELECTRON MICROSCOPES, ANALYSIS OF MAGNETIC FIELD DISTRIBUTION OF SOME TYPES OF MAGNETIC LENSES BY FEM Holčáková R. 1, Marek M. 1, Folvarčný A. 1, Jelen L. 2, Režnar J2 1

VSB-Technical University of Ostrava, FEI, KAT410, Laboratory of Magnetic Measurements and Applications, 17 listopadu 15, Ostrava-Poruba, 70800 Czech Republic; [email protected], [email protected]

2

VÍTKOVICE - Research and Development- Technical Applications a.s., Studentská 6202/17, 70800, Ostrava-Poruba; Czech Republic

The paper deals the basic goals and results of the project specialized in innovation of research in the magnetic lens and chambers of electron microscopes. This project and paper were created by financial support of state budget through the Ministry of Industry and Trade MPO-CR, project n. FRTI1/334. The paper deals with the partial results of the project, concretely the calculating simulation of the distribution magnetic field in the type electron microscope lens. Magnetic field is calculated in different parts of the electron microscope lens: in the structural steel parts of the lens and in the main active vacuum space of the lens. Calculations are solved by finite element method in the software Ansys. The basis of these calculations and simulations are specific geometry of the lens and real magnetic properties of steel. Magnetic properties of steel which is used for construction of magnetic circuit of the lens, were obtained from Laboratory magnetic measurements and applications at the VŠB-TU Ostrava, FEI, KAT410 which was part of the project. Results of calculating simulation of the distribution magnetic field provide a real image of the actual distribution of the magnetic field in the lens and information on potential critical areas which may cause particular defects and inaccuracies of the electron microscope lens.

17

10 WATCHING NANOWIRES GROW Kolíbal M.1, Vystavěl T.2, Novák L.2,Mach J.1, Šikola T.1 1

Institute of Physical Engineering,Brno University of Technology, Technická 2, Brno 616 69; 2 FEI Company, Podnikatelská 6, Brno 612 00

At present semiconductor nanowires are intensively studied for their promising properties in nanoelectronics, photonics, gas and bio-sensing etc. Although the growth process has been well documented already, there are still some issues to be solved, and for this purpose in-situ microscopic experiments are very helpful. In our contribution, we will trace the germanium nanowires (70-300 nm in diameter) growth in-situ from the early stages using scanning electron microscopy. We will show and explain how the changes in the experimental conditions affect the growh process and so the resulting shape of nanowires.

18

11 IN-SITU SEM AND TEM NANOMECHANICAL TESTING OF MATERIALS Lukeš J.1 1

Hysitron Nanomechanical Applications Lab, Technicka 4, 16607 Praha 6.

Nanoindentation has become widly used for materials science research recently. Pressing a dimond tip with an apex radius ≤120nm into the materials and collecting force vs. displacement (depth) data allow us to measure mechanical properties of small volumes of materials or to be more site specific; to test different material phases or very small structures as well as MEMS devices. Obtained indentation curve is a ‘finger print’ of mechanical behavior and is unique for each material. Standard analysis gives us an indentation hardness HIT[Pa] and an elastic reduced modulus [Pa] from material micro- and nano-scale. However, the mechanical properties of nanoscale volumes of material are expected to be enhanced compared to that of the bulk counterparts. What causes the enhanced properties? Is it a fundamental size effect? What role do defects, dislocations or grain boundaries play? All these phenomena can be study with PicoIndenters. PicoIndenters (PI 85 for SEM, PI 95 for TEM, Hysitron, Inc.) provide quantitative nanomechanical testing in conjunction with simultaneous time-synchronized TEM/SEM imaging. Moreover these nanomechanical tests can be combined with a variety of imaging and or analytical techniques (spectroscopy, EBSD). In-situ SEM/TEM nanomechanical testing will be demonstrated on CdS nanosphere compression, nanoindentation of Al grain and nanoindentation of Au thin film.

19

12 VERY LOW ENERGY STEM Müllerová I., Hovorka M., Frank L. Institute of Scientific Instruments ASCR, v.v.i. Královopolská 147, 612 64 Brno, Czech Republic The transmission electron microscope usually operates at primary beam energies (PBE) above 70 keV. Modern scanning electron microscopes with the maximum PBE around 30 keV are also often equipped by a detector of transmitted electrons. We have tested penetrability of free-standing foils in the energy range of the PBE from 10 keV down to nearly 0 eV. Transmissivity of the graphene and of 3 nm and 10 nm foils of gold and carbon, respectively, were measured and series of micrographs obtained vs. PBE. The samples continued to be transparent even at very low energies. The maximum transmissivity (apparently even higher than 100%) was measured for multiple samples at the landing energies around 400 eV [1]. This effect has been explained on the basis of secondary electron (SE) emission released with primary electrons penetrating sufficiently near the exit surface of the foil. Retractable and adjustable version of the detector of transmitted electrons (TE) with a blind central spot was designed and suppression of the SE contribution verified [2]. Graphene flakes were observed under ultrahigh vacuum conditions as well as in a standard SEM environment with similar results. The images formed by TE and by the reflected electrons (RE) have been compared. While RE offered the topographical contrast only, the TE at lowest energies down to units of eV basically exhibit several grey scale levels, which may hypothetically be identified with the number of graphene shells locally overlapped [3]. The experiments were done in SEM devices equipped by the cathode lens [4,5]. [1] Müllerová I. et al., Materials Transactions 51 (2010) 265-270. [2] Müllerová I. et al., in proc. JCNCS 2010, Toyama Japan, 15-18. [3] Müllerová I. et al., IBM J. of Res. and Dev. (2011), in print. [4] Müllerová I. and Frank L., Adv. Imag. & El. Phys. 128 (2003) 309-443. [5] The work is supported by the GAASCR project no: IAA100650902.

20

13 SPECTRAL IMAGING AND LINEAR UNMIXING Krist P. CARL ZEISS spol. s r.o., Radlická 14/3201, 150 00 Praha 5 The dramatic increase in multi-color fluorescence microscopy applications witnessed over the past decade is due, in part, to the significant advances in instrument and detector design as well as the introduction of a vast array of new fluorophores. However, a new problem appeared: so called bleed-through artifacts. Spectral imaging combined with linear unmixing is a highly useful technique that can be used in combination with advanced imaging modalities to untangle fluorescence spectral overlap artifacts in cells and tissues labeled with fluorophores that would be otherwise difficult to separate. An ideal system employs a unique multichannel photomultiplier to gather bands of fluorescence emission light that have been separated into component colors using a diffraction grating. By employing a linear array of detection channels, multiple emission bands are imaged in parallel, thus enabling a selected spectral region to be obtained in a single scan across the specimen (minimizing phototoxicity). Data acquired in such a system can be easily processed even in an online mode. This is especially useful for fast life cell imaging techniques like FRET, FCS, FRAP etc. More information at: http://zeiss-campus.magnet.fsu.edu/index.html http://www.zeiss.cz

21

14 HIGH RESOLUTION LARGE AREA IMAGING OF BIOLOGICAL SAMPLES AND CORRELATIVE MICROSCOPY Gnauck P. Carl Zeiss NTS, Carl Zeis Str. 56, 73447 Oberkochen , Germany We have developed a fast preparation technique that allows a site specific, high resolution investigation the real interface between silicon microstructures and cell tissue at high resolution in a CrossBeam (FIB / FESEM) instrument. Due to the combination of a high resolution field emission SEM for superb imaging capabilities and a high resolution FIB for cutting this technique allows site specific preparation and high resolution investigation of the internal interface at the nm level. The CrossBeam instrument can also be used to do serial sectioning of the sample and provide three dimensional information of the sample at the nm scale. By using very large scan fields I the SEM it is also possible to image large areas at high resolution. By correlating the information obtained in an electron microscope with the fluorescence information that can be obtained in an optical microscope the level of information on a specific sample can be significantly enhanced.

22

15 KONFOKÁLNÍ MIKROSKOPIE LEICA - NOVINKY 2010

Zdeněk Rous Superlativy v konfokální mikroskopii - superrozlišení, supercontinium, supercitlivost. Konfokální mikroskop bez konfokálních irisových šterbin (pinhole)? Leica, Danaher, BalTec, Genetix a kdo se v tom má vyznat?

23

16 NIKON CONFOCAL SYSTEMS AND SUPER RESOLUTION Rozkošný I, Sedlák O, Mack D NIKON s.r.o., Kodaňská 46, 100 10 Praha 10 Nikon presents a new upgrade for Systems C1,/ C1 Si with laser board for 3 or 4 lasers, annual pinhole with 6 sizes, new standard detector with 3 channels, scanning rate max, 2 fps, scanning resolution 2,46 x 2, For systems A1 / A1R / A1 Si / A1R Si are available two scanners type: galvano-scanner, slower with high resolution and resonant scanner, faster with lover resolution. Further 2 laser input ports – with option for assembling from 7 to 9 wavelengths, maximal resolution 4096 x 4096 points, maximal rate A1: 4 fps (512x512); A1R: 30 fps (512x512) – 420 fps (512x32). Hexagonal-pinhole – continuous adjustable from 12 to 256 µm.4 channels PMT detector. 32channels spectral detector (options Si), resolution 2.5/6/10 nm, rate 4 fps with 256 x 256 points resolution. Optional VAAS detector (Virtual Adaptable Aperture System). FAST and DEEP in vivo imaging with A1R MP multiphoton confocal system, compatible with MP femto-second lasers Mai Tai HP DeepSee Chamelenon Vision. Nikon N-STORM (multicolor 2D, 3D: licensed from Harvard-University) Stochastic optical reconstruction microscopy makes use of photo-activated fluorochromes. Nikon N-SIM Microscopy, (multicolor 2D, 3D, TIRF; licensed of UCSF) makes use of structured illumination. 2x higher resolution in comparison with possibilities of optical microscopy. With acquisition rate 0,6 pictures per second, this method is suitable for live cell imaging

24

17 PERKINELMER - LIVE CELL IMAGING AND IMAGE DATA MANAGEMENT Horák R.1 1

PE Systems, s.r.o., výhradní zástupce PerkinElmer pro ČR

PerkinElmer’s live cell imaging expertise and detailed knowledge of your scientific applications means that we can provide the most advanced live cell imaging solutions specific to your needs. The UltraVIEW VoX is the world’s first fully end-to-end spinning disk confocal microscope, from acquisition to analysis. It has been designed to meet the challenges of your live cell imaging research, including the risk of damage from fluorescence illumination and high power laser excitation. Scientists working with images generate massive amounts of data that need to be accessed quickly, analyzed and re-analyzed, shared with colleagues and stored safely. Until now managing that data has been difficult and limiting. Columbus® is the first universal high-volume image data management and analysis system that brings fast, secure access to images from a wide range of sources via the Internet.

25

18 ®

THE NANOWIZARD 3 – THE MOST FLEXIBLE, HIGH RESOLUTION AFM WITH TRUE OPTICAL INTEGRATION Mark Richter, Heiko Haschke, Torsten Jaehnke, Rachel Owen, Gerd Behme JPK Instruments AG, Bouchéstrasse 12, 12435 Berlin, Germany The NanoWizard®3 represents the latest in AFM technology. The new Vortis™ controller series uses the latest FPGA architecture to guarantee highest digital performance. Fast signal acquisition and control, advanced feedback and analysis are key components of a modular and ultra flexible controller. The high-speed data acquisition makes the controller perfect for time resolved force spectroscopy, higher harmonics imaging or high frequency cantilever use. Cantilever calibration by thermal noise method up to 3.25 MHz is unique. HyperDrive™ is a soft sample imaging technique in liquid which provides subnanometer lateral resolution with minimal tip-sample interactions and works with off-the-shelf cantilevers. This is made possible by the new optics and electronics of the NanoWizard®3 AFM head, which gives the lowest noise level in the cantilever deflection detection system available commercially. The NanoWizard®3 maximizes stability, performance and ease of handling for samples in fluid and for full integration with optical microscopy. This enables the simultaneous acquisition of high quality AFM images with optical imaging, under physiological conditions. This is critical as for biological samples, whether cells or molecules, extra information from optical signals can be vital to interpreting the AFM images. This can be fluorescent markers of certain components, for instance, or structural information from optical phase contrast or DIC. The unique DirectOverlay™ software for the JPK NanoWizard® systems uses the tip location to calibrate accurately the optical images and integrate them into the AFM software for direct AFM navigation. In addition, exact, quantitative correlation of AFM and optical features is possible. With the NanoWizard®3, AFM imaging and force spectroscopy can also be combined simultaneously with high end optical techniques such as confocal LSM, FCS, FRET, FRAP or TIRF.

26

19 BIOLOGICAL APPLICATIONS MICROSCOPE MIRA 3 FEG SEM

OF

THE

SCANNING

ELECTRON

Kubíčková K. Tescan, a.s., Libušina třída 21, 623 00 Brno One of the principle goals of TESCAN is to extend applications of TESCAN scanning electron microscopes to the field of biological research. MIRA 3 FEG SEM for high vacuum as well as for low vacuum operations is an ideal tool for reaching excellent results in Biology, Medicine and Food Science. High Resolution Schottky Field Emission SEM supplemented by the optional accessories (STEM adaptor, LVSTD detector, Water Vapour Inlet, Peltier Cooling Stage, Cryo-SEM preparation system) allows observation of morphology, internal microstructure and chemical constitution of biological samples. The accessories offer great opportunity to obtain superb quality images with using all advantages of MIRA 3 FEG SEM.

27

20 CONFOCAL FLUORIMETRY – ANALYTICAL APPROACH Klepetář J.1, Janáček J.1 1

Institute of Physiology of the Academy of Sciences of the Czech Republic, v.v.i. LSCM can be exploited as the fluorimeter for both end-point and kinetic measurements, and for estimations based on fluorescence decay measurements. LSCM combined with quantitative image analysis has very distinct advantages in determinations of substrates, enzymes and antigenes, however the method has some limitations. Approach to the confocal fluorimetry based on analytical chemistry procedures, the quality control and statistical routines included, can bring some benefits. Supported by by grant P108110794 from the Grant Agency of the Academy of Sciences of the Czech Republic,

28

21 ANALYSIS OF NANOPARTICLES IN SOFT ORAL TISSUES Benada O.1, Venclíková Z.2, Joska L.3 1

Institute of Microbiology v.v.i., Academy of Sciences of the Czech Republic, Vídeňská 1083, 142 20 Prague 4, CR; 2 Institute of Dental Research, 1st Faculty of Medicine, Charles University in Prague and General University Hospital, Karlovo náměstí 32, 121 11 Prague 2, CR; 3 Department of Metals and Corrosion Engineering, Institute of Chemical Technology, Technická 5, 166 28 Prague 6, CR. Argyria, characteristic and irreversible bluish-gray pigmentation of a skin and mucous membranes, is generally classified as localized or generalized one. Distinct pigmentation of the oral mucosa in the vicinity of amalgam fillings is often referred to as amalgam tattoos. It is also classified into the category of non-melanine-associated pigmented lesions of the oral cavity. An amalgam is generally considered as the cause of this pigmentation. However, pigmented areas can be also associated with silver-containing corrosion products of dental alloys used for prosthetic restorations. We have shown that formation of soluble silver compounds in the gingival sulcular area or in a crevice between the crown and the cast post- and core reconstruction is possible. Electron microscopy of biopsies revealed numerous submicroscopic electron-dense particles (EDPs) of various size and localization in the tissue. EDPs were found extracellularly as well as intracellularly, mainly in fibroblasts, rarely in macrophages and endothelial cells, but never in the epithelium. Elemental composition of EDPs determined by x-ray microanalysis showed mainly silver in combination with sulfur or selenium or both chalcogens.The results of elemental analysis of silver-containing EDPs were merged together with the excerpts from the patients clinical records. This work was supported by grant NK 7437-3, NR9124/3 (MH, CR), and Institutional Research Concept AV0Z50200510 (AS CR).

29

22 ARTIFACT-REDUCTION STRATEGIES IN BIOLOGICAL PHASE-CONTRAST MICROSCOPY Pelc R.1,3,4, Hostounský Z.2, Kim C.-S.3 1

Inst. Physiol., Acad. Sci., CZ-14200 Prague 4 Stentor Inst., CZ-25301 Hostivice-Palouky 614 3 Warm-Temp. Forest Res. Ctr., Jeju 697-050, Korea 4 JBRI Hi-Tech Develop. Inst., Jeju 697-943, Korea 2

Translucent, non-absorbing cells (so-called phase objects) can only be visualized by microscopic optical contrasting if no chemical staining is applied. Interference microscopy as an ideal mode (no longer commercially available) yields images that are artifact-free. Phase-contrast represents an alternative but images of optically not-too-thin cells suffer from halo/shade-off artifacts. These can be attenuated e.g. by apodization [1] as documented here in pollen grains of Taxus sp. and plant leaf replicas. However, this strategy results in contrast deterioration in optically very thin objects such buccal epithelial cells, seed wings of Pinus sp. (Czech R.) or glycerol-embedded (refractive-indexmatched) trichomes of Elaeagnus sp (Jeju Island). Other setups capable of reducing the imaging artifacts such as American Optical’s “Polanret“ or Zeiss‘ “Interphako“ (neither of them commercially available any longer) are briefly (re)introduced. Supported by NRF/KOSEF, LC06063 and Nikon CZ. [1] Pelc, Hostounský & Otaki (2008) J. Biomed. Opt. 13(5): 054067 A

B

C

A seed wing of Pinus sp. in (A) bright field, (B) conventional and (C) apodized phase contrast (positive type). 300x300 µm

30

23 HOW TO IMPROVE PREPARATION OF SAMPLES FOR IMMUNOGOLD LABELING ON ULTRATHIN SECTIONS? Sobol M.A.1*, Philimonenko V.V. 1, Philimonenko A.A. 1, Janda P. 2, Nebesářová J. 3 , Hozák P. 1 1

Department of Biology of the Cell Nucleus, Institute of Molecular Genetics ASCR, v.v.i., Vídeňská 1083, 142 20 Prague, Czech Republic; *[email protected] 2 J. Heyrovský Institute of Physical Chemistry ASCR, v.v.i., Dolejškova 2155/3, 182 23 Prague, Czech Republic 3 Laboratory of Electron Microscopy, Biology Centre of ASCR - Institute of Parasitology, Branišovská 31, 37005 Ceske Budejovice, Czech Republic We present and discuss how to preserve both the ultrastructure and antigenicity of human cultured interphase and mitotic cells allowing the subsequent immunogold labeling on thin sections. The approach combines HPF and FS followed by resin embedding with some essential recommendations. We compare various HPF cryoprotectants to achieve an improved cell ultrastructure. The cryoprotection method of choice will be discussed. We introduce effective FS procedure of cryofixed cells with a use of glutaraldehyde and water added to acetone to minimize the sample structural distortions. FS protocol followed by LR White embedding favors a preservation of both fine morphology and contrast in interphase cells, but also in mitotic cells that are usually very complicated to preserve. Detailed recommendations concerning preferable protocol will be presented. We report a comparison of several embedding resins as regard to their ability to expose antigens on ultrathin sections for immunolabeling. The resin surface cut using a diamond knife is observed with AFM, and results are statistically analyzed. We conclude that LR White resin can be well combined with HPF/FS for fine ultrastructural immunocytochemistry giving good preservation of both fine ultrastructure and the immunolabeling intensity. Supported by grants from AV ČR (KAN200520704) and MŠMT ČR (LC545, 2B06063).

31

24 MULTIPLE IMMUNOLABELING OF ANTIGENS ON RESIN SECTIONS OF BIOLOGICAL SPECIMENS Hozák P.1,Philimonenko V. 1, Philimonenko A. 1, Nebesářová J. 2, Šlouf M. 3, Halbhuber Z. 4, Krivjanská M. 4, 1

Institute of Molecular Genetics ASCR v.v.i, Biology Centre ASCR v.v.i.; 3 Institute of Macromolecular Chemistry ASCR v.v.i.; 4 Central European Biosystems, s.r.o. 2

Simultaneous detection of antigens by means of indirect immunolabeling provides valuable information about their localization in cellular compartments and their possible interactions in macromolecular complexes. However, ultrastructural immunodetection is limited to sensitive simultaneous labeling of only two antigens. For the purpose of overcoming this limitation, we prepared a set of novel nanoparticles with the size around 10 nm and good properties for the use as labels in electron microscopy. These nanoparticles can be distinguished from commercial spherical gold nanoparticles by their shape or by elemental composition. The nanoparticles were successfully conjugated to various secondary antibodies and probed for the ability of immunolabeling on ultrathin resin sections of cells. Consequently, we created functional sets of our new conjugates and antibodies with commercial labels to be used for simultaneous detection of up to five antigens. We demostrate an example of successful fivetuple immunolabeling for transmission electron microscopy, where labels are easily distinguishable from each other on the basis of their shape. Moreover, the use of some combinations including quantum dots would allow targets detection in correlative light and electron microscopy. Acknowledgement: KAN KAN200520704, MŠMT LC06063, institutional grant AV0Z50520514.

32

POSTERY (uspořádáno podle jména prezentujícího autora)

33

1 BILATERAL CHANGES OF IL-6 PROTEIN AND ITS RECEPTORS IN DORSAL ROOT GANGLIA FOLLOWING UNILATERAL CHRONIC COMPRESSION INDRY Brázda V.1,2, Jagelská B. E.1, Dubový P.2, Svíženská I. 2, Klusáková I.2 1

Institute of Biophysics Academy of Sciences Czech Republic, Brno, Czech Republic 2 Department of Anatomy, Division of Neuroanatomy, Medical Faculty, Masaryk University, Brno, Czech Republic Many experimental studies have documented important role of cytokines and immune cells during different types of neuropathy, and their potential to induce or facilitate neuropathic pain. Interleukin 6 (IL-6) is a proinflammatory cytokine involved as a key signal molecule in the neuronal and immune responses to nerve injury. A biological effect of IL-6 is mediated by two signaltransducing receptors: IL-6 receptor (IL-6R) and glycoprotein 130 (gp-130). We have investigated immunohistochemical localization of IL-6, IL-6R and gp-130 in the ipsilateral and contralateral L4-L5 dorsal root ganglia (DRG) following spinal nerve ligature (SNL) in comparison with those from intact and sham-operated rats. Our results indicate that the satellite glial cells of DRG are important source of IL-6 protein. Moreover, these cells also display changes of corresponding receptors in both ipsi- and contralateral DRG following unilateral nerve injury inducing neuropathic pain. The results suggested a possibility of indirect biological effect of IL-6 to neuronal activity mediated by corresponding receptors of the satellite glial cells.

34

2 SEM CHARACTERIZATION OF PD NANOPARTICLES PREPARED BY REVERSE MICELLE TECHNIQUE Černohorský O.1,2, Žďánský K.2 1

Department of Physical Electronics, Czech Technical University, V Holesovickach 2, Prague, CR; 2 Institute of Photonics and Electronics, Academy of Sciences of the Czech Republic, Chaberska 57, Prague, CR Layer of Pd nanoparticles on InP can be used as a hydrogen sensor. Generally, energetic barrier called Schottky barrier is formed on the interface between metal and semiconductor (i.e. Pd and InP in our case). Hydrogen atoms dissociated by Pd nanoparticles lowers this Schottky barrier and we can then measure hydrogen concentration by electric current. We prepared Pd nanoparticles in reverse micelles with AOT as a surfactant and isooctane as a solvent. The layers of these particles on InP were prepared by electrophoretic deposition. This method consist in the acceleration of Pd nanoparticles from their colloid solution in the direction of the InP substrate in the presence of electric field. Deposited Pd nanoparticles formed aggregates on the substrate. Their size depended on the parameters of electrophoretic deposition. For characterization of these layers, I-V measurement, C-V measurement, secondary-ion mass spectroscopy, AFM, SEM, and measurement of response of the structure to the presence of hydrogen were used.

35

3 POINT SPREAD FUNCTION DEGRADATION IN THICK TISSUES Chernyavskiy O.1, Kubínová L. 1 1

Institute of Physiology ASCR, v.v.i..

Although image acquisition by a confocal microscope provides true 3D imaging, it is often affected by a number of factors deteriorating the overall image quality. Postprocessing algorithms, such as deconvolution, applied to raw data can significantly improve the image quality, e.g. resolution and signalto-noise ratio. For example, deconvolution can improve volume estimation results [1]. The experimental point spread function (PSF) is usually measured from confocal microscopic 3D images of microbeads having subresolution size, embedded in medium of the same refractive index (RI) as the used objective immersion. Different biological tissues have different optical properties (e.g., RI and transparency). This can cause considerable degradation of the PSF, especially in deeper layers of the specimen. PSF changes in different depths of a biological specimen can be measured in different ways [2, 3]. In the present study we evaluated the depth dependence of experimental PSF in thick tissue sections. We measured PSF directly from microbeads located in different depths of the specimen. Such measurements provided more precise information not only for data quantification, but also can be used for testing different deconvolution algorithms. This study was supported by the Academy of Sciences of the Czech Republic (Institutional Research Concepts No. AV0Z50110509), the Grant Agency of the Czech Republic (grant 304/09/0733), and Ministry of Education, Youth and Sports of the Czech Republic (research program LC06063 and ME09010). References [1] F. Difato, F. Mazzone, S. Scaglione, M. Fato, F. Beltrame, L. Kubínová, J. Janáček, P. Ramoino, G. Vicidomini, A. Diaspro, Microsc. Res. Tech. 64 (2004) 151. [2] M. von Tiedemann, Microsc. Res. Tech. 69 (2006) 10. [3] A. Diaspro et al., Applied Optics 41 (2002) 685.

36

4 X-RAY MICROSCOPY WITH MEDIPIX2 Dammer J.1, Weyda F.2, Sopko V.1 1

Institute of Experimental and Applied Physics, Czech Technical University in Prague, Horska 3a/22, CZ-12800 Prague 2, Czech Republic 2 Biology Centre of the Academy of Sciences of the Czech Republic v. v. i., Institute of Entomology, Branisovska 31, CZ-37005 Ceske Budejovice, Czech Republic We describe the recently developed radiographic apparatus, equipped with Medipix2 semiconductor pixel detector. The detector is used as an imager that counts individual photons of ionizing radiation, emitted by an X-ray tube (micro- or nano-focus FeinFocus). Thanks to the wide dynamic range of the Medipix2 detector and its high spatial resolution better than 1µm, the setup is particularly suitable for radiographic imaging of small biological samples. Along with the description of the apparatus we provide examples of the iodine contrast agent (Optiray) in vivo application as a tracer in various model organisms. The iodine contrast agent increases the absorption of X-rays and this leads to better resolution of some internal structures of organisms, especially the various cavities, pores, etc. Similar results presents another tracer, lanthanum nitrate. Microradiographic imaging also helps detect organisms living in a not visible environment non-destructively, visualize the dynamic internal biological processes and also to resolve some details of their body (morphology). Tiny phytophagous insects and other arthropods are an ideal object for our studies. This work was realized in frame of the CERN Medipix Collaboration and was supported in part by the Research Grant Collaboration of the Czech Republic with CERN No. 1P04LA211, by the Fundamental Research Center Project LC06041 and the Research Programs 6840770029 and 6840770040 and Grant No. 2B06005 of the Ministry of Education, Youth and Sports of the Czech Republic.

37

5 COALESCENCE IN QUIESCENT IMMISCIBLE POLYMER BLENDS: ELUCIDATION OF MATRIX PHASE STATE INFLUENCE USING SEM Dimzoski B., Šlouf M., Fortelný I., Mikešová J. Institute of Macromolecular Chemistry, Academy of Sciences of the Czech Republic, Heyrovsky Sq. 2, 162 06 Prague 6, Czech Republic. Introduction Coalescence of dispersed droplets in quiescent polymer blends at temperatures higher than melting temperature is driven by the tendency to achieve morphology with minimum interfacial area. Elucidation of matrix phase state influence on the morphology evolution for immiscible polymer blends with crystalline and amorphous matrix was studied by SEM. Experimental Immiscible polymer blends prepared in identical processing conditions, with 15 wt% of EPR as dispersed phase and PP (crystalline) or PS (amorphous) matrix, were annealed in quiescent state at 180 oC. Morphology evolution was observed after certain annealing time intervals (until 60 min) using scanning electron microscope VEGA TS 5135 (Tescan). Micrographs were evaluated regarding the changes of the size of dispersed droplets using specialized image analyses program (NIS-Elements, LIM). Results and Conclusion Image analyses of acquired SEM micrographs provided a quantitative structure descriptor – average equivalent diameter – which was employed in description of the morphology evolution. Bigger EPR particle size observed immediately after the melt mixing of PS/EPR blend could be attributed to the higher interfacial tension in comparison with PP/EPR blend. Slower growth rate of the EPR particle size with the annealing time was perceived for polymer blend with PP matrix, although the zero shear viscosity of PP at 180 oC is relatively lower than the corresponding one for PS, increasing the mobility of the dispersed phase. This observation suggested that the matrix phase state could have impact on phase structure: the droplets could break up during the matrix crystallization processes. Acknowledgement: GAČR P106/11/1069

38

6 FLUORESCENCE LIFETIME IMAGING AND SPECTROSCOPIC CHARACTERIZATION OF FIREFLY LUCIFERIN IN LAMPYRIS NOCTILUCA Fischerová J.1,2, CHorvát D.1,2 1

International Laser Centre, 2Comenius University, Bratislava, Slovakia.

Bioluminescence in Lampyris noctiluca is the light emission that appears as a result of catalytic reaction of luciferin with luciferase enzyme with the presence of ATP and oxygen. Luciferin, in addition to its bioluminescent character, is also a fluorescent pigment. In this study we utilized an advanced microscopy system combining fluorescence microscopy, spectroscopy and fluorescence lifetime detection that have been recently developed in our laboratory, to characterize spatial and spectral properties of luciferin directly in the intact animal. Spatial distribution of Luciferin in Lampyris noctiluca was imaged either by custom fluorescence macro-imaging setup, or by confocal laser scanning microscope (Zeiss LSM 510 META) with 2-photon excitation and fluorescence lifetime imaging subsystem (Becker-Hickl). For spectroscopic characterization of both animal and purified Luciferin we used absorption, fluorescence and time-resolved spectroscopy setups. The results indicate that spatial distribution of Luciferin fluorescence in Lampyris noctiluca is not constrained to its luminescent organs. We also demonstrated that spectrally resolved fluorescence lifetime microscopy provides a synergic effect with great potential for gathering detailed information on photophysical behaviour of luciferin inside the intact insect, helping to understand the structure and function of firefly bioluminescent system. Authors acknowledge funding from the project NanoNet (ITMS: 26240120010) under the OP Research and Development of the European fund for regional development and grant VEGA No. 1/0530/09.

39

7 STUDY OF HUMAN EMBRYONIC STEM CELLS WITH SCANNING ELECTRON MICROSCOPE Flodrová E.1,2, Neděla V.1, Sedláčková M.3, Hampl A.3 1

Institute of Scientific Instruments of the ASCR, v.v.i, Department of Electrotechnology, The Faculty of Electrical Engineering and Communication, Brno University of Technology, 3 Department of Histology and Embryology, The Faculty of Medicine, Masaryk University. 2

Potential differentiation to all the types of cells in the organism and the ability of self-renewal, makes human embryonic stem cells (hESCs) an important research subject of modern medicine. There are great expectations on them especially for their ability to cure a range of so far incurable diseases. Deeper understanding many diseases is expected too. Using scanning electron microscopy (SEM) the specifics of the surface of hESCs can be displayed and then studied. Cells have to be fixed and dried for SEM observations. To prevent a modification of the surface structure, it is necessary to treat cells together with the substrate (glass, plastic) to which they adhere. Particularly the drying process is the critical operation for this type of samples. It leads to erosion of integrity of individual colonies. Finding the suitable methodology allowing showing the samples of undifferentiated hESCs, cultured on glass or plastic disc with a supporting layer of mouse embryonic fibroblasts is very important then. [1]Inoué, T., Osaka, T.: J. Electron Microscopy 38 (1989), p. 246– 249. [2]Nation, J.L.: Stain Tech. 58 (1983), p. 347 – 351. [3]This work was supported by the Academy of Sciences of the Czech Republic, Grant No. GAP 102/10/1410

40

8 FIB INDUCED DAMAGE EXAMINED WITH TEM Frank L.1, Mikmeková Š.1, Matsuda K.2, Watanabe K.3, Ikeno S.2, Müllerová I.1 1

Institute of Scientific Instruments of the ASCR, v.v.i., Czech Republic; Graduate School of Science and Engineering for Research, University of Toyama, Japan; 3 Graduate School of Science and Engineering for Education, University of Toyama, Japan. 2

One of the most important applications of the focused ion beam (FIB) technique is preparation of samples for observation in the transmission electron microscope (TEM). However, the FIB can induce reconfiguration or destruction of the specimen structure and in crystalline materials it can create an amorphous layer on the surface. Thickness of the damage caused by the ion bombardment was estimated by Monte Carlo simulations using the SRIM code and the calculated data have been compared with experimental values obtained by means of the TEM. Experimental Results Range by SRIM Calculation

70 damage depth (nm)

60 50 40 30 20 10 0 0

20

40

60

80

angle of incidence (°)

Calculated and measured damage depth as a function of the angle of incidence.

41

9 PROSPECTS OF THE SCANNING MICROSCOPY IN MATERIALS SCIENCE

LOW

ENERGY

ELECTRON

Frank L.1, Mikmeková Š.1, Konvalina I.1, Müllerová I.1 ,Hovorka M.1 1

Institute of Scientific Instruments of the ASCR, v.v.i., Královopolská 147, 612 64 Brno, Czech Republic Employment of the scanning low energy electron microscopy (SLEEM) has been slowly making its way into the field of materials science, hampered not by limitations in the technique but rather by the relative scarcity of these instruments in research institutes and laboratories. SLEEM is a method especially useful for observing the grains in polycrystalline materials thanks to its high spatial resolution and fast acquisition of data when comparing with the EBSD method, and to the possibility of observing spacious bulk samples. The cathode lens (CL) mode in the SEM enables us to detect slow but not only slow, high angle scattered electrons that carry significant crystallographic contrast based on the electron channeling. Good prospects of the SLEEM in materials science follow from many opportunities the method provides not only in fundamental research but also in analytic study of materials important for practice.

42

10 VISUALIZATION OF PROTEIN DISTRIBUTION INSIDE POROUS CHROMATOGRAPHIC ADSORBENT BY TRANSMISSION ELECTRON MICROSCOPY Gramblička M.1, Uhrík B.2, Polakovič M.1, Sulová Z.2, Rusnák A.2 1

Faculty of Chemical and Food Technology, Slovak University of Technology in Bratislava; 2 Institute of Molecular Physiology and Genetics, Slovak Academy of Sciences This study presents an attempt to investigate the internal porous structure of a chromatographic adsorbent by transmission electron microscopy. The adsorbent was a strong cation exchanger FractoGel EMD SE Hicap (Merck, Germany). It is a crosslinked polymethacrylate resin with a mean pore diameter of 24 nm (determined by the dextran size exclusion). The adsorbent was primarily designed for purification of monoclonal antibodies and represents a modern generation of materials with a very high adsorption capacity (> 150 mg IgG per ml of resin) enabled by the surface grafting technology. Multiple methods of sample preparation were tested, involving various combinations of fixation by glutaraldehyde and staining by osmium and uranyl acetate. Two types of samples were investigated: the adsorbent devoid of protein (a blank test) and the adsorbent saturated by human IgG at very favourable binding conditions (100 mM acetate buffer at pH 4.5, adsorbed protein over 150 mg/ml). After the fixation and staining, the sample was polymerized into a block of epoxy resin (Durcupan) and cut by an ultramicrotome into slices of the thickness of 58 nm. The results indicated that a suitable choice of the staining method is crucial. The sample stained by osmium followed by uranyl acetate exhibited a sharp contrast of the porous structure, whose dimensions were in agreement with the pore size range determined by the dextran size exclusion. This method might enable a precise estimation of pore tortuosity and connectivity. Acknowledgements: Grants APVV-0084-07, APVV-VVCE-0064-07, VEGA 1/0655/09

43

11 ELECTRON MICROSCOPY OF POROUS SEMICONDUCTORS Grym J.1, Nohavica D.1, Jarchovský Z.1, Piksová K.2 1 2

Institute of Photonics and Electronics AS CR; Faculty of Nuclear Sciences and Physical Engineering, CTU Prague

We report on the SEM characterization of porous networks in III-V semiconductors. The main goal was to achieve homogeneous nucleation and gain control over the density of pores in the nucleation layer and its branching below this layer. The control is essential for the application in heteroepitaxial growth with a high lattice mismatch between the substrate and the deposited layer or in the preparation of composite materials on the nanoscale. The pore etching was carried out in an electrochemical cell containing a fluoride-iodide aqueous electrolyte (H2O-HF-KI) for GaAs and a chloride electrolyte (H2O-HCl) for InP using a configuration equivalent to four electrodes. A home-made potentiostat/galvanostat was computer-controlled and allowed to register all process variables. (100)-oriented GaAs:Si and InP:Sn substrates with a carrier concentration ranging from 1x1017 to 2x1018 cm-3 were used for the pore preparation. We discuss the influence of the (i) electrolyte type and anion concentration; (ii) substrate, its crystallographic orientation, dopant concentration; (iii) current/voltage regime; and (iv) illumination on the pore formation observed in Phillips XL30 ESEM and JEOL JSM 7500F scanning electron microscopes. We also present some data from AFM measurements and Nomarski-contrast optical microscopy. The work was supported by the project P108/10/0253 of the Czech Science Foundation.

44

12 SCANNING ELECTRON MICROSCOPY CHARACTERISTICS OF THE (PHARYNGODONIDAE)

FOCUSED GENUS

ON THE KEY CITHARINIELLA

Hodová I., Koubková B., Baruš V. Department of Botany and Zoology, Faculty of Science, Masaryk University, Kotlářská 2, 611 37 Brno, Czech Republic. Three nematode species of Cithariniella (Pharyngodonidae), C. citharini, C. khalili, and C. gonzalesi, were recorded from the recta of squeaker (Mochokidae: Siluriformes) and citharinid (Citharinidae: Characiformes) fishes from Senegal, West Africa. Morphological characteristics obtained by scanning electron microscopy (form of oral aperture and cephalic papillae, presence of lateral alae, distribution and form of cloacal papillae, simple or paired papillae on tail of males, eggs with numerous long filaments on each pole in females) correspond well to the generic diagnosis and represent species differences. The shape and size of the cephalic papillae and lips were identified as a new determination feature. C. gonzalesi is reported for the first time from Senegal and its host, Paradistichodus dimidiatus, represents a new host record. Morphometrical features of these small nematode species observed under light microscope are very similar and therefore the methods of SEM were used for detecting valid morphological characteristics for differetiation.

45

13 MAPPING OF DOPANTS IN SILICON BY INJECTION OF ELECTRONS Hovorka M., Konvalina I., Frank L. Institute of Scientific Instruments of the ASCR, v.v.i., Brno, Czech Republic Dopants in silicon based structures locally modify the secondary electron emission, revealing in this way their distribution over the sample. For probing the doped structures usually the electron beam is used at energies around 1 keV. However, the very low landing energy range has proven itself an efficient tool for mapping dopants in semiconductors [1, 2]. We have focused on planar p-type structures of various dopant densities, embedded in an n-type substrate. The samples were observed in UHV electron microscope with the cathode lens. Imaging by means of secondary electrons and its quantifiability was verified and the method was extended to very low energies. At few keV and hundreds of eV the detected signal is composed of a mixture of secondary and backscattered electrons. Contrast between p-type patterns and n-type substrate reaches its maximum around 1 keV where the contribution of secondary electrons dominates, and then the contrast gradually decreases. At the tens and units of eV a dynamical contrast connected to the charge injection via primary electrons is observed. P-type patterns charge up and then totally reflect the incident electron beam. Contrast formation depends on whether the reflected beam hits the detector or its central bore and is influenced by electron dose and other parameters. The dopant concentration in p-type patterns can be quantified via contrast dependency on the electron dose. In vicinity of a charged area on the surface the trajectories of electrons landing at low energy are significantly modified. We have simulated the electron trajectories inside the cathode lens for such case and compared the calculated and measured intensity distributions on the screen. [1] Frank L., Müllerová I., Ultramicroscopy 106 (2005), p. 28-36. [2] Hovorka et al., Proceedings of LEEM/PEEM 6 (2006), Trieste, p. 110.

46

14 3D ANALYSIS OF CAPILLARIES Janáček J.1, Kubínová L.1, Chernyavskyi O.1, Eržen I.2, Mao X.W.3 1

Institute of Physiology ASCR; 2Univerzity of Ljubljana; 3Loma Linda Univerzity.

Confocal images of capillaries can be used for 3D vizualization and analysis of the capillary network. Capillaries in the rat brain were stained by whole-body perfusion. Capillaries in rat or human muscles were stained by multiple immunostaining. The capillaries were detected, after appropriate image preprocessing, by thresholding and by tracing the skeletonized 3D image. The results were corrected using desktop virtual reality environment. The 3D models were used for visualization and estimation of length density, tortuosity, anisotropy and average length of capillaries. In the skeletal muscle the characteristics of capillary network in the vicinity of individual fibers can be measured. The values can serve for characterization of capillary bed in different tissues, of patholgies or of effects of experimental treatment. The work was supported by grant ME 09010. Čebašek, V., Eržen, I., Vyhnal, A., Janáček, J., Ribarič, S., Kubínová, L.: The estimation error of skeletal muscle capillary supply is significantly reduced by 3D method. Microvascular Research 79(1): 40-46, 2010. Mao, X.W., Favre, C.J., Fike, J.R., Kubínová, L., Anderson, E., CampbellBeachler, M., Jones, T., Smith, A., Rightnar, S., Nelson, G.A.: High-LET Radiation-Induced Response of Microvessels in the Hippocampus. Radiation Research 173(4): 486-493, 2010.

47

15 CLOSTRIDIUM TYROBUTYRICUM PREVENTES DSS INDUCED COLITIS AND REGULATES IL-18 EXPRESSION IN THE MICE COLON – QUANTITATIVE IMAGE ANALYSIS CONTRIBUTION J. Klepetarb, T. Hudcovica, J. Kolinskab*, , V. Erbanc, H. Kozakovaa a

Institute of Microbiology of the Academy of Sciences of the Czech Republic, v.v.i. b Institute of Physiology of the Academy of Sciences of the Czech Republic, v.v.i. c Food Research Institute, Prague, Czech Republic Clostridium tyrobutyricum prevented dextran sulphate(DSS) induced colitis in mice. Various parameters were evaluated and IL-18 in mucosa of the colon was visualized by process with primary and secondary antibody labeled with Cy-3 fluorophore. Laser scanning confocal microscopy (Leica SPE ) and the quantitative image analysis supplemented by statistical evaluation of results were applied to the sections with visualized IL-18. The results of quantitative image analysis are in excellent agreement with other histological, clinical and biochemical findings at immunocompetent mice strain BALB/c, thus fully support protective effect of C.tyrobutyricum against DSS induced colitis. The results with immunodeficient mice strain SCID support certain considerations based on experimental results. The results of quantitative image analysis conclusively show the potentiality of confocal immunofluorimetry. supported by grants 303/08/0367 and 303/09/0449 of the Czech Foundation of the Czech Republic, grants 2B06155 and ME10017 of the Ministry of Education, Youth and Sports, by grant S5002005720 from the Grant Agency of the Academy of Sciences of the Czech Republic, and by Institutional Research Concept AV0Z50200510 and AV0Z50110509.

48

16 IMAGE CONTRASTS IN THE SCANNING ELECTRON MICROSCOPY Konvalina I., Hovorka M., Mikmeková Š., Müllerová I. Institute of Scientific Instruments of the ASCR, v. v. i. The image contrast in the scanning electron microscope (SEM) is governed not only by interaction of the primary electrons with the specimen but also by setup of the microscope. The same specimen can exhibit different contrasts in various SEMs and even in one microscope different detectors show the specimen differently. Position of the detector in the specimen chamber and spatial distributions of the electrostatic and magnetic fields around the specimen and detector play in the contrast formation the key role [1]. When interpreting the image contrasts we have to consider all instrument parameters that influence the transport of signal electrons to the detector. Several examples are presented. The field strength of the cathode lens [2] can collimate high angle backscattered electrons to the detector, which are capable of producing micrographs of high contrasts of the local crystallinity. Magnetic field of the objective lens (OL) either penetrates toward the specimen surface or remains confined inside OL. Signal electrons influenced by the magnetic field move along a spiral trajectory and a part of them is retracted toward the specimen (so called "bottle effect"), which leads to losses of contrast. The Everhart-Thornley detector of secondary electrons (SE) placed below OL collects about 30 % of the emitted SEs. Variations in the working distance influence the collection of SEs leaving the specimen under larger polar angles. [1] Müllerová I., Konvalina I., Journal of Microscopy. 2009, vol. 236, pp. 203-210. [2] Müllerová I., Frank L., Advances in Imaging and Electron Physics. 2003, vol. 128, pp. 309-443. [3] Supported by the GAASCR (IAA100650902).

49

17 STRUCTURAL CHARACTERIZATION OF TiOx NANOTUBE ARRAYS AND PREPARATION OF TEST STRUCTURES FOR ELECTRICAL MEASUREMENT Križanová Z.1, Vávra I.1, Bahayeu S.2 1

Institute of Electrical Engineering, Slovak Academy of Sciences, Dúbravská cesta 9, 841 04, Bratislava, Slovakia 2 Physical-Technical Institute, National Academy of Sciences of Belarus, 1/3 Kuprevich St., Minsk 220141, Belarus Titanium oxide thin films have received a great deal of interest because of an importatnt number of applications, such as antireflection coatings, microelectronic devices, devices for the purification and treatment of water and air, or as gas sensors (H2, NO, NO2, CO). A variety of method have been used for the preparation of TiOx. The same applies to relatively recenly discovered highly ordered TiOx nanotube arrays. It is very interesting material having especially large internal surface. It has been well established that their properties, structure and phase composition depend on deposition conditions. This poster shows a study of ordered TiOx nanotube arrays using Transmission Electron Microscope (TEM) and Scanning Electon Microscope (SEM). The initially fabricated nanotube arrays are usually almost amorpous and crystallizing with a hight temperature annealing. They have simillar pore diameters, wall thicknes and length. And for the purpose of their electrical measurement, we prepared them by Foucused Ion Beam microscope – Quanta 3D 200i. The FIB equipment (Quanta 3D 200i) was purchased within the project of the structural funds of the European Union entitled: „Centre of excellence for new technologies in electrical engineering“, ITMS code 26240120011

50

18 STRUCTURAL CHARACTERISATION OF POLYELECTROLYTE CAPSULES BY SCANNING ELECTRON MICROSCOPY Krzyžánek V.1, Keller U.1, Sporenberg N.2, Guddorf J.2, Reichelt R.1, Schönhoff M.2 1

Inst. Med. Physics & Biophysics; 2Inst. Phys. Chem., University of Münster, Robert-Koch-Str. 31, D-48149 Münster, Germany Polyelectrolyte hollow capsules are a promising material that can be used for example as a drug delivery system in medical applications [1]. They can be prepared from polyelectrolytes on silica particles by using the layer-bylayer self assembly method [2] and, consequently, dissolving the silica core with dilute hydrofluoric acid. The presented capsules are formed of 5 bilayers of the PAH/PSS polyelectrolytes with a diameter of ~400 nm. Scanning electron microscopy was employed to investigate the structure details of the capsule wall arising from the core dissolution process. Because the capsules are damaged during their drying in air due to the surface tension forces, the freeze-drying techniques developed and used mainly for investigations of biological samples were used to prevent the intact shape of the capsules. Capsules were rapidly frozen into liquid ethane, subsequently freeze-dried at vacuum, and finally rotary coated with Pt/Ir/C for high-resolution imaging. This technique allowed for the first time to observe the capsules at high resolution in their intact shape and find, e.g., holes within size ≥10 nm in their capsule wall, thus, helping to improve the capsule production. In addition, backscattered electron micrographs of the capsules before dissolving the silica core allow rough estimation of the wall thickness that is important for the next quantitative characterisations of the capsules. [1] [2] [3]

AL Becker et al: Small 6 (2010) 1836. RP Choudhury and M Schönhoff: J. Chem. Phys. 127 (2007) 234702. Supported by DFG grant RE 728/11-2.

51

19 COMPARISON OF METHODS FOR CHLOROPLAST ESTIMATION – DISECTOR AND PROFILE COUNTING

NUMBER

Kubínová Z.1, Lhotáková Z.1, Kubínová L.2, Albrechtová J.1 1

Charles University in Prague, Faculty of Science, Department of Experimental Plant Biology, Viničná 5, 128 44 Prague 2, Czech Republic; 2 Institute of Physiology, AS CR, v.v.i., Vídeňská 1083, 142 20 Prague 4, Czech Republic Chloroplasts are organelles of plant cells, where photosynthetic processes take place. Chloroplast number within a mesophyll cell can be influenced by environmental factors and therefore it is frequently examined parameter. We compared two methods for estimation of chloroplast number per cell – the disector method, which is based on 3D unbiased sampling probe (Sterio, 1984; Gundersen, 1986) and one of the commonly used methods counting chloroplast profiles in thin physical (2D) sections of the leaf. Needles collected from Norway spruce trees were stored frozen before processing. Cross-sections were cut off in a systematic uniform randomly chosen positions along the needle. Their images were captured by a Leica SP2 AOBS confocal microscope using 10x objective. Systematically sampled series of optical sections were acquired with higher resolution using 63x objective. The series were then used for counting chloroplasts. We found that the profile counting method underestimated the number of chloroplasts per cell significantly; the obtained values were about ten times lower than values obtained by the unbiased disector method. The optical disector method enables unbiased estimation of particle number in 3D. We applied disector method for chloroplast counting in studies of effect of elevated CO2 concentration on the anatomical structure of sun and shade Norway spruce needles. Supported by the Ministry of Education, Youth and Sports CR (LC 06063), Academy of Sciences CR (AV0Z50110509), and by the Grant Agency CR (P501/10/0340).

52

20 EVALUATION OF MICROSCOPY IMAGES IN MATERIAL SCIENCE Kunka I.1, Starý V.1 1

Faculty of Mechanical Engineering of Czech Technical Univ. in Prague, Dept. of Mat. Eng., Karlovo nám. 13, CZ-121 35 Prague 2, Czech Republic

Studying the morphology of the material (the structure in terms of layout components, including its size, number and spatial distribution) is usually taken by microscopy. The observed structure corresponds to the used magnification (millimeter, micrometer, nanometer, or sub nanometer scale - atomic), which we obtain using a light or electron microscope. In any case we get a digital picture of the structure that we have to process quantitatively or qualitatively. Helping hands are modern and intuitive software, which we must calibrate with the used device and check due to the applicable standards. When using any type of microscope, unfortunately, we are not always able to obtain high image quality. Important role plays the noise, uneven distribution of the intensity of the background or of the objects. Very often it`s necessary to eliminate unwanted objects which does not belong to the image, or are not included to our measurement aim. Processing problems of these "problematic" images is the main goal of this work.

53

21 MEASUREMENT OF Pd NANOPARTICLE DIAMETERS USING BSE IMAGING IN FESEM Langhans J.1,3 ,Nebesářová J.1,2, Dean J.L.1,3 , Vancová M.1, Šlouf M.4 , Pavlová E.4 1

Biology Centre of ASCR, České Budějovice, Faculty of Science, Charles University in Prague, Praha, 3 Faculty of Science, University of South Bohemia, České Budějovice, 4 Institute of Macromolecular Chemistry of ASCR, Praha 2

The possibilities of using a field emission scanning electron microscope (FESEM) equipped with an Autrata improved yttrium aluminum garnet (YAG) detector[1] of back-scattered electrons (BSE) for to determine the size of Pd nanoparticles were examined in this study. Pd nanoparticles with expected approximate sizes of 14, 10 and 6 nm were prepared according to our recently developed procedure [2], which was based on a general protocol described earlier by Turkewich [3].The diameters of Pd nanoparticles were determined from digitally recorded FESEM micrographs and were compared with the diameters obtained with a transmission electron microscope (TEM). On the basis of obtained results, we can conclude: 1/ The measurement of the size distribution Pd nanoparticles with Autrata YAG detector in FESEM is possible, although its accuracy is influenced by a number of factors, e.g. accelerating voltage, size, magnification as well as image processing. 2/ Pd nanoparticles are less stable under the primary electron beam and yield lower signal in comparison with Au nanoparticles. Therefore, the size of the smallest 6nm Pd nanoparticles could not be reliably measured. [1] R. Autrata R., EMSA Bull 1992, 22, 54-58. [2] M. Slouf, E. Pavlova, M.S. Bhardwaj, J. Plestil, H. Onderkova, A.A.Philimonenko, P. Hozak, Materials Letters 2010, submitted. [3] J. Turkevich, G. Kim, Science, 1970, 169, 873-879. [4] Acknowledgement: KAN200520704, Z60220518

54

22 INNOVATIVE RESEARCH IN ELECTRON MICROSCOPES, MAGNETIC PROPERTIES OF STEELS AND ALLOYS FOR MAGNETIC LENSES AND CHAMBERS Marek M. 1, Folvarčný A. 1, Holčáková R. 1, Jelen L. 2, Režnar J2 1

VSB-Technical University of Ostrava, FEI, KAT410, Laboratory of Magnetic Measurements and Applications, 17 listopadu 15, Ostrava-Poruba, 70800 Czech Republic; [email protected]; 2 VÍTKOVICE - Research and Development- Technical Applications a.s., Studentská 6202/17, 70800, Ostrava-Poruba; Czech Republic

The paper deals with the basic goals and partial results of the project which focuses on innovation of research in the magnetic lenses and chambers of electron microscopes. This project and paper were created by financial support of state budget through the Ministry of Industry and Trade MPO-CR, project n. FR-TI1/334. The paper contains the analysis of magnetic properties of the types steels and alloys which are designed for construction of lens and chamber of the electron microscopes. This paper presents results of measuring magnetic properties of the materials: pure ferrum - Behanit, alloy FeNi, types construction steel for chambers and same other special types. This paper presents the BH stationary hysteresis characteristics and magnetizing characteristics and next dynamic characteristics. The self measurement of magnetic properties was carried out in the Laboratory Magnetic Measurements and Applications at the VSB-TU Ostrava, FEI, KAT410. For measuring were used the precision digitally controlled measuring systems for DC and AC hysteresis characteristics. Results and methods of this work bringing new optimization possibilities for the construction of the electromagnetic lens for electron microscopes.

55

23 STEREOSCOPIC SCANNING ELECTRON MICROGRAPHS OF HEAD STRUCTURES OF PARASITIC VERTEBRATE NEMATODE Mašová Š.1, Wandrol P.2, Baruš V.1 1

Department of Botany and Zoology, Faculty of Science, Masaryk University, Kotlářská 2, 611 37 Brno, Czech Republic; 2 FEI Czech Republic s. r. o., Podnikatelská 6, 612 00 Brno, Czech Republic. This poster describes ascaridid nematode species Dujardinascaris madagascariensis Chabaud et Caballero, 1966, which was obtained by parasitological dissection from the stomach of a perished Nile Crocodile at Lake Turkana. Nematodes have been studied by scanning electron microscope to determine and redescribe their external morphology. For SEM, specimens were prepared by the standard method. Beside classic scanning electron micrographs we used also stereomicrographs for morphologic observations of nematode head structures. Two micrographs taken at moderately different viewing angles were used for the stereoscopic imaging. The optimal angle difference is 4-6 degrees. Obtained images make use of the parallel viewing method or they can be joined into stereoscopic pictures (also known as 3D anaglyphs). Anaglyphs are pictures where the red and blue channels have been split and then reassembled so that the image appears three-dimensional when viewed through 3D glasses with red and blue lenses. Three-dimensional images are very useful for better visualization of feature arrangement and their spatial distribution comparing to the ordinary scanning electron micrographs. They also bring more realistic insight on nematode morphology. Acknowledgements: This study was supported by the Long-Term Research Plan MSM 0021622416 funded by the Ministry of Education, Youth and Sports of the Czech Republic, also by the GACR Grant No. 526/09/H025 and by the FEI Czech Republic s.r.o.

56

24 MIKROSKOPICKÁ DOKUMENTACE POŠKOZENÍ STRUKTURY DŘEVA BĚŽNĚ POUŽÍVANÝMI BIOCIDNÍMI PROSTŘEDKY Michalcová A.1, Kučerová I1, Drábková K.1 1

Ústav chemické technologie restaurování památek, VŠCHT Praha, Technická 5, Praha 6, 166 28

Historické dřevo bývá běžně ošetřováno ochrannými přípravky na bázi anorganických solí, které mají fungicidní účinek nebo působí jako retardéry hoření, přestože není stále dostatek informací o jejich možném negativním účinku na dřevo z hlediska jeho životnosti. Jako fungicidy na bázi anorganických solí se dnes užívají směsi solí a/nebo oxidů B, Cu, Cr a Zn. Mezi látky s retardační účinností na bázi anorganických látek patří široké spektrum anorganických chemikálií, např. amonné soli, sloučeniny bóru, halogenidy aj. V této práci je popsán vliv síranu měďnatého a kyseliny borité na poškození struktury dřeva. Vzorky dřeva byly impregnovány 5 % roztoky studovaných chemikálií a následně vystaveny podmínkám urychleného stárnutí (expozice při teplotě 70 °C – suché stárnutí, a při teplotě 70 °C relativní vlhkosti 80 % vlhké stárnutí, po dobu 30 dnů). Vzorky s obsahem síranu měďnatého vykazovaly odlišnosti v poškození v závislosti na podmínkách urychleného stárnutí: U vzorků exponovaných v podmínkách vlhkého stárnutí se výrazně změnil makroskopický i mikroskopický vzhled struktury dřeva. Výsledky ukázaly, že dochází především ke korozi celulózy. Vzorky exponované v podmínkách suchého stárnutí jsou charakterizovány změnami ve struktuře ligninu. Některé vzorky s obsahem kyseliny borité z mikroskopického hlediska vykazovaly oddělování vláken z buněčné stěny dřeva v případě vzorků exponovaných v podmínkách vlhkého stárnutí v důsledku změn ve struktuře celulózy. Tyto výsledky ukazují, že obě běžně používané anorganické soli poškozují strukturu dřeva, proto je jejich aplikace na historické dřevo nevhodná. Tato práce vznikla v rámci řešení projektu MSM 6046137302 a MŠMT CR COST OC08042.

57

25 NON-RIGID REGISTRATION OF CLSM IMAGES OF PHYSICAL SECTIONS WITH DISCONTINUOUS DEFORMATIONS BY MULTI-LABEL OPTIMIZATION Michálek J.1, Čapek M.1,2, Kubínová L.1 1 2

Institute of Physiology AS CR, v.v.i., Prague Faculty of Biomedical Engineering, CTU, Kladno, Czech Republic

When biological specimens are cut into physical sections for threedimensional (3D) imaging by confocal laser scanning microscopy (CLSM), the slices may get distorted or ruptured. For subsequent 3D reconstruction, images from different physical sections need to be spatially aligned by optimization of a functional composed of a data fidelity term evaluating similarity between the reference and target images, and a regularization term enforcing transformation smoothness. Regularization term evaluating the total variation (TV), which enables the registration algorithm to account for discontinuities in the slice deformation (ruptures), while enforcing smoothness on continuously deformed regions, was proposed previously. The functional with the TV regularization was optimized using a graph-cut (GC) based iterative solution. However, GC may generate visible registration artifacts, which impair the 3D reconstruction. We present an alternative, multi-label, TV optimization algorithm which in the examined examples prevents the artifacts produced by GC. The algorithm is slower that GC, but can be sped up several times when implemented in a multiprocessor computing environment. Besides, for image pairs with uneven brightness distribution, we introduce a reformulation of the TV-based registration, in which intensity-based data term is replaced by comparison of salient features in the reference and target images quantified by local image entropies. Supported by grants of MŠMT ČR MSM6840770012, LC06063, ME09010, GAČR 102/08/0691, 304/09/0733, the Academy of Sciences of the Czech Republic (Institutional Research Concept No. AV0Z50110509).

58

26 APPLICATION IN LABORATORY FOR ELECTRON MICROSCOPY Mika F., Hanzlíková R., Rek A. ÚPT AV ČR, Královopolská 147, 612 64 Brno. The Laboratory of Electron Microscopy (LEM) of the ISI AS CR v.v.i. is a highly respected research institute equipped with unique instruments. Its staff consists of experts in development and application of electron microscope instrumentation and methodology. Currently, LEM offers imaging and analysis of solid surfaces at high spatial resolution using numerous unique techniques. Very low energy SEM exhibits plenty of novel contrast mechanisms reveal electronic and crystallinic structures of the sample. Instrumentation accessories allowing this imaging method are available for multiple instruments, including our ultra-high-vacuum system. Environmental SEM enables investigation of living substances and materials under elevated pressure of surrounding gas up to 3000 Pa. For examination of nanomaterials with ultimate spatial resolution, we use the cold field emission electron source available in the JEOL JSM 6700F SEM. This tool is capable of imaging with verified resolution of 1 nm at the electron energy of 15 keV. The microscope is equipped with the energy dispersive analyzer of X-rays for chemical microanalysis. LEM provides standard sample preparation techniques such as sputtering and evaporation of surface coatings, ion beam thinning and exact cutting. Complementary light optical imaging is possible with the confocal microscope Olympus LEXT 3100. Further information on our equipment and available techniques may be obtained from http://www.lem.isibrno.cz.

59

27 IN VIVO TWO-PHOTON EXCITATION IMAGING OF NEURONAL NETWORKS

FLUORESCENCE

CALCIUM

Novák O.1, Syka J.1 1

Department of Auditory Neuroscience, Institute of Experimental Medicine, Academy of Sciences of the Czech Republic, v.v.i.

Information processing in the mammalian cortex is among the most complex activities that can be found in nature. To understand such complex processes, it is necessary to study the interactions between neurons in cortical networks, which can be achieved with the aid of real-time imaging with sufficient resolution. Neuronal action potentials can be observed indirectly, through associated changes in the intracellular calcium concentration. Changes in the calcium concentration can be monitored with a Ca2+dependent fluorescent probe. Biological tissue, however, strongly absorbs and scatters light, making it impossible to obtain sufficient resolution in vivo using conventional methods of fluorescence microscopy. These problems, however, may be eliminated by non-linear, in our case two-photon, laser scanning microscopy. Thanks to the phenomenon using the effect of higher order intensity, the focal plane is very clearly defined by the objective itself, and any additional pinhole is not necessary. Also, much longer excitation wave-lengths can be used, which significantly reduce absorption and scattering in the studied tissue. Because of these effects, the neuronal structures can be studied within the sufficient range of depths. In the Department of Auditory Neuroscience, Institute of Experimental Medicine, AS CR, v.v.i. in Prague, we use two-photon laser scanning microscopy for studying neuronal activity in the auditory cortex of anaesthetized young rats in an in vivo configuration. The studied group of neurons is stained with the MCBL method (Stosiek et al., 2003). The configuration of our set-up was completed and the optimal adjustment of the experimental protocol was finalized only recently.

60

28 CORRELATED OPTICAL MICROSCOPE/RAMAN MICROSCOPE/SCANNING ELECTRON MICROSCOPE STUDY OF SELFASSEMBLED GOLD NANOROD ARRAYS Novotný F.1, Proška J.1, Procházka M.2 1

Faculty of Nuclear Sciences and Physical Engineering, Czech Technical University in Prague 2 Faculty of Mathematics and Physics, Charles University in Prague The excitation of localized surface plasmons at metallic nanoparticles (MNPs) by light, denoted as localized surface plasmon resonance (LSPR), is phenomenon which attracts a great deal of attention in both theoretical and experimental research. Moreover, during past decade a number of differrent MNPs shapes was synthesized and those method refined to a point, where high-yield samples of non-spherical metallic MNPs can be obtained. The unique electro-optical properties of such MNPs induced number of applications; spanning from fundametal studies of colloidal systems to aplication in biological systems as a biocompatible and addressable carrier. We have prepared high quality voluminous domains of several hundred cubic micrometers of gold nanorod (GNR) supracrystals through the surfactant driven phase separation. As the GNRs have interesting plasmonic properties, composites based on arranged GNRs are supposed to be very promising materials with respect to plasmon coupling and interactions with light. We present experimental study of correlated optical microscope/SEM measurement showing different optical response of GNR arrays in dependence of the GNR orientation inside the self-assembled array together with surface enhanced raman scattering measurement. Acknowledgement: This research was KAN401220801.

supported

by

61

GAAV,

Czech

Republic,

project

29 BETTER TO SEE ONCE THAN HEAR A HUNDRED TIMES - CONFOCAL MICROSCOPY OF ENZYMES IN PLANT CARBOHYDRATE METABOLISM Ovečka M.1,3, Li J.1, Ezquer I.1, Bahaji A.1,2, Baroja-Fernández E.1, José Muñoz F.1, Almagro G.1, Montero M.1, Hidalgo M.1, Teresa Sesma M.1, Pozueta-Romero J.1 1

Instituto de Agrobiotecnología Mutiloabeti, Nafarroa, Spain; Iden Biotechnology S.L., Pamplona, Nafarroa, Spain; 3 Institute of Botany, SAS, Bratislava, Slovakia 2

Metabolism of carbohydrates in plants is highly regulated and connected with various biological processes in response to the physiological needs of the cell. Starch is the main storage carbohydrate in plants. Identification and understanding of all factors that are involved in the starch biosynthetic process is important for many aspects of agricultural and economic human needs. Although starch biosynthesis occurs in the plastids of plant cells, some critical reactions take place in the cytosol. Thus, subcellular localization studies are important to confirm where main enzymes and substrates are located, synthesized and/or utilized. To address this problem, we have produced and characterized transgenic potato and Arabidopsis plants constitutively expressing a translationally fused green fluroescent protein (GFP)-encoding genes under the control of the cauliflower mosaic virus 35S promoter. By using this tool and a confocal microscope we localized ADP-sugar pyrophosphatase (StASPP-GFP), hydrolysing the starch precursor molecule, sucrose synthase (SuSy-GFP), converting sucrose and a nucleoside diphosphates, soluble starch synthase IV (SSIV-GFP) and granule-bound starch synthase (GBSS1GFP). We proved cytosolic localization of EcASPP and SuSy, plastidial localization in the stroma of StASPP, uniform localization of GBSS1 in starch granules and specific localization of SSIV in regions associated with the edges of the starch granules. Results confirmed compartmentalization of multi-step starch metabolic pathway allowing storage and reuse of energy in the form of complex starch grains. Partly supported by VEGA 2/0200/10.

62

30 SEM CHACTERIZATION OF METAL NANOPARTICLES DEPOSITED ON SEMICONDUCTORS Piksová K.1, Koštejn M.1, Grym J.2, Müller M.1 1

Department of Physical Electronics, Faculty of Nuclear Sciences and Physical Engineering, Czech Technical University in Prague, Břehová 7,115 19 Prague 1; 2 Institute of Photonics and Electronics AS CR, Chaberska 57, 18251 Prague 8

We report on the SEM characterization of metal nanoparticles deposited by electrophoresis on semiconductor wafers. The motivation is prepared high quality nanolayers for diodes. Schottky barriers with layers of nanosized metal particles on semiconductor wafers can be used as hydrogen sensors or radiation detector. Colloids of metal nanoparticles stabilized by AOT reverse micelles in isooctane were prepared. The colloids contain nanoparticles with the size distribution of 5-10 nm in diameter. The morphology of the deposited layers was observed by JEOL JSM 7500F scanning microscope. This work has been supported by the Grant Agency of the Academy of Science of the Czech Republic, project KAN401220801 and the Czech Science Foundation grant 102/09/1037

63

31 MAPPING OF THE LOCAL DENSITY OF ELECTRON STATES BY VERY LOW ENERGY ELECTRON REFLECTIVITY Pokorná Z., Frank L. Institute of Scientific Instruments AS CR, v.v.i. Královopolská 147, 612 64 Brno, Czech Republic Local density of states is an important characteristic of solids, in particular the crystals. One way of probing the local density of states is to measure the reflectivity of very low energy electrons (units to tens of eV) from the sample surface. The reflectivity should be inversely proportional to the local density of electronic states coupled to the impinging electron wave [1]. A cathode lens-equipped [2] scanning electron microscope can effectively map the reflectivity of very low energy electrons, maintaining a high lateral resolution. The cathode lens is a zero working distance electrostatic lens where the specimen is at a negative potential and serves as the cathode. This arrangement allows decelerating the incident electrons arbitrarily. The density of states, probed in direction perpendicular to the surface, varies with orientation of the crystal face. A series of demonstration experiments was carried out on several differently oriented single crystal aluminum samples and on a sample of the Highly Ordered Pyrolytic Graphite (HOPG). The reflectivity was compared to theoretical calculations [3,4] and the influence of surface cleanliness and sample preparation techniques is discussed. [5] [1] V.N. Strocov, H.I. Starnberg, Phys. Rev. B 52 (1995) 8759. [2] I. Müllerová, L. Frank, Adv. Imaging and Electron Phys. 128 (2003) 309. [3] E. Krasovskii, W. Schattke, Phys. Rev. B 56 (1997) 12874. [4] E. Krasovskii, Phys. Rev. B 70 (2004) 245322. [5] Supported by the CSF under the grant no. P108/11/270.

64

32 PREPARATION OF SUBSTRATES FOR SURFACE-ENHANCED RAMAN SPECTROSCOPY: SELF-ASSEMBLY AND BIOTEMPLATING Proška J.1, Novotný F.1, Štolcová L. 1, Berger J.1, Procházka M.2 1

Faculty of Nuclear Sciences and Physical Engineering, Czech Technical University in Prague; 2 Faculty of Mathematics and Physics, Institute of Physics, Charles University in Prague The phenomenon of surface-enhanced Raman scattering (SERS) attracts great interest due to amplifying molecular Raman signal with many orders of magnitude. Common types of substrates with rugged surfaces are metal colloids, or metallized microparticles, galvanically "etched" electrodes, and lithographically prepared periodic arrays of distinct shapes. Appropriate biotemplates can be used for preparation of less sterically limited configuration of metal nanoparticles. Such materials often yield number of 'hot-spots' via plasmonic coupling of closely spaced nanoparticles. To this purpose we have used butterfly and cicada wings. Microsphere self-assembly, shadow lithography, electroless silver plating, relief casting, and gold, silver and copper magnetron sputtering were used to prepare the effective substrates for SERS. Modified bio-templates and microparticle assemblies were successfully tested and high quality SERS spectra were obtained. This work was supported partially by grants of GAAV KAN401220801, Ministry of Education, Youth and Sports, project COST OC09038, and MŠMT MSM0021620835 (to M. Procházka)

65

33 CHALLENGES OF HIGH-PRESSURE FREEZING AND FREEZE SUBSTITUTION TECHNIQUES IN PRESERVATION OF CHLOROPLAST ULTRASTRUCTURE OF FAGUS SYLVATICA LEAVES AND PICEA ABIES NEEDLES Radochová B.1, Nebesářová J.2, Lhotáková Z.3, Čapek M.1, Kubínová L.1, Albrechtová J.3 1

Institute of Physiology, Academy of Sciences of the Czech Republic, Vídeňská 1083, 142 20 Praha 4; 2 Institute of Parasitology, Academy of Sciences of the Czech Republic, Branišovská 31, 370 05 České Budějovice; 3 Charles University in Prague, Faculty of Science, Department of Experimental Plant Biology, Viničná 5, 128 44 Praha 2. High-pressure freezing (HPF) and freeze substitution (FS) are powerful techniques enabling us to study cell ultrastructure close to its living state. Samples up to thickness of about 200 µm can be preserved, which is sufficient for leaves of most plant species. However, in the case of plant tissues, HPF and FS techniques have proved to be quite challenging, mainly due to large water content in plant cell vacuoles and therefore the risk of ice crystal formation leading to damage of cell structures. In the present study we investigated effect of two different cryoprotectants (dextran and hexadecen) as well as three different FS-media (2% OsO4 + 0,25% glutaraldehyde in anhydrous acetone, 2% + 0,25% glutaraldehyde in anhydrous acetone with addition of 1% H2O and 2% OsO4 + 0,2% uranyl acetate in anhydrous acetone) on the chloroplast ultrastructure of Fagus sylvatica leaves and Picea abies needles. HPF and FS proved to be challenging, especially in the case of Picea abies needles, where the cells were seriously damaged, regardless of the type of cryoprotectant or the type of FS-media used. Therefore we need to test other types of cryoprotectants and make adjustment of the entire HPF and FS process. Acknowledgements: This work was supported by the Grant Agency of the Czech Republic (project no. P501/10/0340).

66

34 CHANGES OF SURFACE PROPERTIES OF TiO2 Říhová Z. 1, Starý V. 1, Bačáková L. 2 ¨ 1

Faculty of Mechanical Engineering of Czech Technical University in Prague, Department of Materials Engineering., Karlovo náměstí 13,121 35 Prague 2, Czech Republic 2 Institute of Physiology of AS CR, Vídeňská 1083, 141 20 Prague 4, Czech Republic The work concerns about the morfology and stability of biocompatible nanostructured TiO2 surfaces which are deposited on Ti6Al4V alloy. The morphology of the deposited layers was observed by scanning electron microscope (SEM).The layer was rated by elemental surface analysis. The morphology of the layer influences the contact angel. Stability is significant for provision and innovation of biocompatibility coating. We studied the changes of contact angle before and after sterilization and during some time on the samples in different environments. Samples were located in four different environments: distilled water, air, physiology solution, dry air (in desiccator). Because during the first measurement the samples can be influenced by UV light, there were during the second measurement situated in dark place. The surface energy was calculated from measured contact angles of distilled water and ethylenglycol according to the model Owens – Wendt. From the tests results follows the TiO2 coatings of all samples in the darkness were time stable. Comparing with first measurement, when samples were not placed in the dark, proved the influence of the surface photoactivity on the contact angle size. The changes of contact angles before and after sterilization were not indicate. Acknowledgment: This work was supported by the project KAN101120701 of Grant agency of Academy of Science of Czech Republic. We would like thank to Ing. Jirků from firm Medin, a.s., Nové Město na Moravě for providing the samples.

67

35 ROLE OF VINCULIN DURING THE MOUSE SPERMATOGENESIS Rohožková J., Venit T., Hozák P. Dept. of biology of the cell nucleus, Academy of Science Czech Republic, Vídeňská 1083, Praha 142 20 Vinculin is a member of proteins described as molecules responsible to sense the mechanical properties of the extracellular environment, and to act as a regulator of mechanical stress in addition to its function as a mechanocoupling protein. Vinculin is the main component of the focal adhesions establishing cell-cell and cell-matrix interaction. Disrupting of vinculin leads to deregulation of mentioned interactions and increased cell migration in a 3D environment. This, in turn, is considered prerequisite for the development of malignant tumors Till present the presence of vinculin was described in U2OS on the ultrastructural level. Except prove of the presence of this molecule in interphase cell we were interested in the localization of vinculin during the meiosis. Using the immunofluorescence method we have described presence of vinculin on the mouse testes cryo-sections (on the basement t of Ductus seminiferi) and on the single meiotic cell spreads. Vinculin is localized as a diffused signal all over the nucleus in preleptotene/leptotene cell and colocalizes with SCP3 protein. This protein is main component of synaptonemal complex (SC) created between homologous paired chromosomes. During the meiotic division cell undergoes extensive changes in shape, size and movement. Thy cytoskeleton, which comprises actin, microtubules and intermediate filaments, is believed to function in these cellular events. What is the concrete role of vinculin, the actin binding protein, still remains unclear.

68

36 BIOSYNTHESIS OF NANOPARTICLES USING DIATOMS MICROSCOPICAL CHARACTERIZATION Schröfel A.1, Kratošová G.1, Vávra I.2, Bohunická M.3 1

Nanotechnology Centre, VŠB-Technical University in Ostrava; Institute of Electrical Engineering, Slovak Academy of Sciences; 3 Faculty of Science, University of South Bohemia 2

Synthesis of gold nanoparticles, EPS-gold and silica-gold bionanocomposites by biologically driven processes employing two diatom strains (Navicula atomus, Diadesmis gallica) is described by means of microscopy techniques. Transmission electron microscopy (TEM) and electron diffraction analysis (SAED) revealed a qualitative presence of gold nanoparticles in the experimental solutions of the diatom culture mixed with tetrachloroaureate. Scanning electron microscopy (SEM) and TEM showed that the nanoparticles were associated with the diatom frustules and extracellular polysaccharides (EPS) excreted by the diatom cells. Optical microscopy techniques described overall changes in the diatom cells and also confirm presence of the nanoparticles (purple color caused by surface plasmon resonance) on the surface of the cells or the cell debris. This study confirmed microscopic techniques as proper approach for description of metal nanoparticles in biological systems. In particular, combination of high resolution SEM microscope with cryo-system is easy and very powerful method. Moreover, chemical nature of analyzed materials can be confirmed and proved either by SAED or combination of detectors (YAG, LEI).

69

37 MORPHOLOGY STUDY OF LARVAL INSTARS BY DUAL BEAM MICROSCOPE Semelbauer M.1, Šmatko V.2 , Kozánek M.1, Vávra I.2 1

Institute of Zoology, SAS, Dúbravská cesta 9, 84506 Bratislava Slovakia Institute of Electrical Engineering, SAS, Dúbravská cesta 9, 84506 Bratislava Slovakia

2

We used a dual beam microscope to describe the external morphology of larval instars of Minettia longipennis (Fabricius, 1794) (Diptera, Lauxaniidae). Larvae were killed in hot water (40°C) and dehydrated through 80, 90, 99,5% ethanol and hexamethyldisilazane and finally coated with gold. The larvae of many Diptera species store spherical inorganic concretions in Malpighi tubes. Third-instar larva of M. longipennis has notably enlarged the distal parts of the Malpighi tubes, which serve as storage for mineralised concretions. For a detailed study of cirri the cross-section was performed by ion beam milling. The chitin thickness on the larval body surface is low (compared with that of adult) and therefore there is a danger that during observation (by ebeam or ion beam) the hole in the larva will be opened and the mineralised concretions will contaminate the microscope stage. The FIB equipment (Quanta 3D 200i) was purchased within the project of the structural funds of the European Union entitled: „Centre of excellence for new technologies in electrical engineering“, ITMS code 26240120011

70

38 CHARACTERIZATION OF ONE-STEP AND SEQUENTIALLY IRRADIATED UHMWPE FOR TOTAL JOINT REPLACEMENTS Šlouf M.1, Baldrian J.1, Kotek J.1, Lednický F.1, Šandová J.1, Hromádková J.1, Fencl J.2, Bouda T.3, Janigová I.4 1

Institute of Macromolecular Chemistry AS CR, Praha Beznoska Ltd., Kladno 3 Czech Technical University, Praha 4 Polymer Institute SAS, Bratislava 2

Ultra-high molecular weight polyethylene (UHMWPE) is the most common bearing material for total joint replacements (TJR) of big human joints, such as hip, knee, shoulder, ankle etc. In order to increase the lifetime of TJR, the UHMWPE is crosslinked by ionizing radiation, such as gamma rays or accelerated electrons (gamma or e-beam irradiation). The crosslinking increases UHMWPE wear resistance and, consequently, the lifetime of TJR as the wear is regarded as the main cause of TJR failures. A standard UHMWPE crosslinking includes a one-step irradiation followed by remelting (heating above Tm = 140 oC). Recently, a group of investigators suggested that the sequential irradiation, which consists in three-step irradiation with one third of the final radiation dose followed by three annealings (heating below Tm) should improve the mechanical properties in comparison with the standard treatment. In this work, we compared six UHMWPE samples: non-irradiated, irradiated in a standard way and sequentially irradiated, which were both remelted and/or annealed. The structure was characterized by a number of microscopic (LM, SEM, TEM), diffraction (SAXS, WAXS), spectroscopic (IR, ESR) and thermal (DSC) methods. The properties were assessed by small-punch (SPT), microhardness (MHv) and wear testing. All experiments were in quite a good agreement, showing that the improvement of the properties in sequentially irradiated UHMWPE is questionable. Acknowledgement: MŠMT 2B06096, TAČR TA01011406.

71

39 PREPARATION AND CHARACTERIZATION OF NANOPARTICLES FOR MULTIPLE IMMUNOLABELLING Šlouf M.1, Pavlova E.1, Vlková H.1, Hrubý M.1, Hromádková J.1, Popelková D.1, Lapčíková M.1, Šloufová I.2, Novotný F.3, Nebesářová J.4, Philimonenko A.5, Philimonenko V.5, Hozák P.5 1 Institute of Macromolecular Chemistry AS CR, Praha 2 Faculty of Science, Charles University, Praha 3 Czech Technical University, Praha 4 Biology Centre AS CR, České Budějovice 5 Institute of Molecular Genetics AS CR, Praha In biology, gold nanoparticles with size 5-15 nm can be conjugated with antibodies and used for immunolabelling. The term immunolabelling means that the antibody, which is attached to the gold nanoparticle by hydrophobic, electrostatic or covalent interaction, can bind to a specific molecule in a carefully prepared microscopic specimen. Subsequently, the specific molecule in the specimen can be detected in an electron microscope, which localizes the nanoparticle (high signal due to high electron density of Au) on the surface of the biological sample (lower signal due to the lighter elements such as C, H, N, O...). Current state-of-the-art in the field of immunolabelling is to detect two molecules simultaneously. This is given by the fact that the commercially available gold nanoparticles in the required size range are spherical, with certain width of distribution. Here we introduce several other nanoparticles, such as Pd, Ag and Co3O4 nanospheres, Pd nanocubes, AgAu core-shell nanoparticles and Au nanorods, which can be conjugated to antibodies and used for multiple immunolabelling, i.e. for the simultaneous microscopic detection of more than two molecules. Acknowledgement: KAN200520704, GACR P205/10/0348 and P208/10/0941, MSM 0021620857.

72

40 ULTRASTRUCTURAL CHANGES IN HUMAN NEURAL CELLS AFTER INFECTION WITH TICK-BORNE ENCEPHALITIS VIRUS Tesařová M. Faculty of Science, University of South Bohemia, České Budějovice, Czech republic. Human cells of neuronal origin represent an excellent tool for the investigation of neuropathogenesis of TBE. The maturation, replication process of tick-borne encephalitis virus (TBEV) and ultrastructural changes induced by infection in the neuroblasts cell line (UKF-NB-4) was studied by electron microscopy. I compared electron microscopical aspects (appearance) of TEM images of neuroblasts cells prepared by (1) conventional chemical fixation, resin-embedding and sectionig; (2) rapid freezing of cell monolayers at high pressure and sectioning of freeze substituted samples. The most interesting fact, however, is that vitrification preserves the cell in close to native state, whereas chemical fixation and dehydration can not take place without extensive intra- and intermolecular cross-linking and aggregation. The appearance of the cytoplasm and nucleoplasm of neuroblasts cells were different in conditions (1) and (2). The excellent ultrastructure of the cytoplasmic and nuclear membranes and organels of neuroblasts cells processed by (2) confirmed the potentional of the method for preservation of cellular fine structures. The infection of neuroblastoma cells was associated with number of major morphological changes, including proliferation of membranes of the rough endoplasmic reticulum and rearrangement of cytoskeletal structures. This report also describes scanning electron microscope study of the surface of neuroblasts cells.

73

41 MICROSTRUCTURE OF TINB ALLOY SURFACE Tolde Z., Anisimov E., Starý V., Pešlová F. Czech Technical University in Prague, Fac. of Mech. Engn., Dept. of Mater. Engn., Karlovo nam. 13, Praha 2, 121 35, Czech Republic Biomaterials for implants are widely used at present. It has been estimated that 90% of the human population above the age of 40 suffer from joint degenerative diseases as arthritis, with causes pain or loss of joint function. In order to maintain the quality of patient’ life it is often necessary to replace the joint with an artificial one. The recent trend in research and development of Ti-alloys for biomedical applications is to develop low-rigidity β-type Ti-alloys, containing non-toxic and non-allergenic elements (Nb, Ta, Zr etc.) with excellent mechanical properties and workability. Coupled with the absence of the reported toxicity of Nb and Ti, Ti-Nb alloys can be used as biomaterials. Moreover these elements in general act as β-Ti-alloy stabilizers. In view of the above considerations, intensive work has been carried out on β-type Ti-alloys with various chemical compositions based on the Ti-Nb binary system. This contribution shows the microscopic observation of the TiNb alloy surface and evaluation of present phases. It also shows the specific possibilities of the behavior of oxide layer.

74

42 MORPHOLOGY AND MECHANICAL PROPERTIES OF PP/COC BLENDS Vacková T1, Šlouf M.1, Nevoralová, M.1 1

Institute of Macromolecular Chemistry, Academy of Sciences of the Czech Republic, Heyrovsky Sq. 2, 162 06 Prague 6, Czech Republic

The aim of this study was to improve mechanical properties of polypropylene / cycloolefin copolymer (PP/COC) blends by processinginduced formation of long COC fibres. We focused our attention on finding processing conditions yielding PP/COC fibrous morphology in a well-defined, reproducible way. A number of PP/COC blends were prepared by both compression moulding and injection moulding (IM). Phase morphology of the resulting PP/COC blends was characterized by means of scanning electron microscopy (SEM), whereas neat polymers were characterized by rheological measurements. The longest COC fibres were achieved in the injection moulded PP/COC blends with compositions 75/25 and 70/30 wt %. Elastic modulus and yield strength of all blends were measured as functions of the blend composition using an Instron tensile tester; statistically significant improvement of the yield strength due to fibrous morphology was proved. Moreover, two different models were applied in the analysis of mechanical properties: (i) the equivalent box model for isotropic blends and (ii) the HalpinTsai model for long fibre composites. In all PP/COC blends prepared by IM, the COC fibres were oriented in the processing direction, as documented by SEM micrographs, and acted as a reinforcing component, as evidenced by stress-strain measurements. This work was supported through Grant 106/09/P272 awarded by the Grant Agency of the Czech Republic.

75

43 TEM CHARACTERIZATION OF of GaP/ZnO2 NANORODS PREPARED ON SINGLE CRYSTALLINE GaP(111) Vávra I., Križanová Z., Hasenöhrl S., Novák J. Institute of Electrical Engineering, Slovak Academy of Sciences, Dúbravská cesta 9, 841 04, Bratislava, Slovakia Nanorods of semiconducting materials epitaxially grown on a semiconductor surface have great potential for the development of new kinds of electronic components for 3D integration. Because of special geometry of the specimen and very small rod diameter, it is necessary to use a special procedure for the TEM specimen preparation. GaP nanorods were prepared by MOCVD technology on the GaP(111) surface. Au nanoparticles (diameter of 30 nm) were used as seeds to catalyze the nanorods growth. After this the surface was covered by ZnO2 (thickness of 10 nm) by rf sputtering technology. Because of a very high fragility of the nanorods, it was not possible to use any standard TEM specimen preparation technology. Therefore, the TEM sample was prepared by Ga ion beam milling of the reverse side of GaP substrate. Only for the final milling the Ar ion beam was used. By TEM investigation we confirmed that the nanorods are epitaxially grown. The detailed TEM analysis revealed a high density of planar structural defects (inside the the rods) The ZnO2 film uniformly covered the rods and also the GaP surface. The ZnO2 is polycrystalline with mean grain size of 10 nm. The FIB equipment (Quanta 3D 200i) was purchased within the project of the structural funds of the European Union entitled: „Centre of excellence for new technologies in electrical engineering“, ITMS code 26240120011

76

Rejstřík autorů Nepodtrženě označené přednášky/podtrženě postery

Albrechtová J. 19 Albrechtová J. 33 Almagro G. 29 Anger M. 3 Anisimov E. 41 Bačáková L. 34 Bahaji A. 29 Bahayeu S. 17 Bakardjieva S. 7 Baldrian J. 38 Baroja-Fernández E. 29 Baruš V. 12, 23 Behme G. 18 Benada O. 21 Berger J. 32 Bohunická M. 36 Bouda T. 38 Brázda V. 1 Čapek M. 25, 33 Černohorský O. 2 Chernyavskiy O. 3, 14 CHorvát D. 6 Dammer J. 4 Dean J.L. 21 Dellvile R. 2 Dimzoski B. 5 Dlabáček Z. 8 Drábková K. 24 Dřínek V. 5 Dubový P. 1 Erban V. 15 Eržen I. 14 Ezquer I. 29 Fencl J. 38

Fischerová J. 6 Flodrová E. 7 Folvarčný A. 9, 22 Fortelný I. 5 Frank L. 12, 8, 9, 13, 31 Ge Y. 8 Gemperle A. 8 Gemperlová J. 8 Gnauck P. 14 Gramblička M. 10 Grym J. 11, 30 Guddorf J. 18 Halbhuber Z. 24 Hampl A. 7 Hannula S.-P. 8 Hanzlíková R. 26 Haschke H. 18 Hasenöhrl S. 43 Hidalgo M. 29 Hodová I. 12 Holčáková R. 9, 22 Horák R. 17 HostounskÝ Z. 22 Hovorka M. 12, 9, 13, 16 Hozák P. 23, 24, 35, 39 Hromádková J. 38, 39 Hrubý M. 39 Hudcovic T. 15 Ikeno S. 8 Jaehnke T. 18 Jagelská B. E. 1 Janáček J. 20, 14 Janda P. 23 Janičkovič D. 6 Janigová I. 38

Jarchovský Z. 11 Jarošová M. 5 Jelen L. 9, 22 Muñoz J. F. 29 Joska L. 21 Keller U. 18 Kim C.-S. 22 Kiselová M. 7 Klementová M. 5 Klepetář J. 20, 15 Klusáková I. 1 Kolíbal M. 10 Kolinska J. 15 Konvalina I. 9, 13, 16 Koštejn M. 30 Kotek J. 38 Koubková B. 12 Kozakova H. 15 Kozánek M. 37 Kratošová G. 36 Krist P. 13 Krivjanská M. 24 Križanová Z. 17, 43 Krzyžánek V. 18 Kubíčková K. 19 Kubínová L. 3, 14, 19, 25, 33 Kubínová Z. 19 Kučerová I. 24 Kunka I. 20 Langhans J. 21 Lapčíková M. 39 Lednický F. 1, 38 Lhotáková Z. 19, 33 Li J. 29 Lukášková Z. 4 Lukeš J. 11 Mach J. 10 Mack D. 16 Mao X.W. 14 Marek M. 9, 22 Mašová Š. 23 Maťko I. 6 Matsuda K. 8 Michalcová A. 24 Michálek J. 25 Mika F. 26 Mikešová J. 5 Mikmeková Š. 8, 9, 16 Montero M. 29 Müller M. 30

Müllerová I. 12, 8, 9, 16 Nebesářová J. 23, 24, 21, 33, 39 Neděla V. 7 Nevoralová M. 42 Nohavica D. 11 Novák J. 43 Novák L. 10, Novák O. 27 Novotný F. 28, 32, 39 Ovečka M. 29 Owen R. 18 Palatinus L. 5 Pavlová E. 21, 39 Pelc R. 22 Pešlová F. 41 Petříček V. 5 Philimonenko A. 23, 24, 39 Philimonenko V. 23, 24, 39 Piksová K. Piksová K. 11, 30 PIlch J. 2 Piluso P. 7 Pokorná Z. 31 Polakovič M. 10 Popelková D. 39 Pospiech M. 4 Pozueta-Romero J. 29 Procházka M. 28, 32 Proška J. 28, 32 Radochová B. 33 Randulová Z. 4 Reichelt R. 18 Rek A. 26 Řezáčová 4 Režnar J. 9, 22 Richter M. 18 Říhová Z. 34 Rohožková J. 35 Rous Z. 15 Rozkošný I.16 Rusnák A. 10 Šandová J. 38 Schönhoff M. 18 Schröfel A. 36 Sedláčková M. 7 Sedlák O. 16 Semelbauer M. 37 Šikola T. 10 Šittner P. 2 Šlouf M. 24, 5, 21, 38, 39, 42 78

Šloufová I. 39 Šmatko V. 37 Sobol M.A. 23 Sopko V. 4 Sporenberg N. 18 Starý V. 20, 34, 41 Štolcová L. 32 Sulová Z. 10 Švec P. 6 Švec SR. P. 6 Svíženská I. 1 Syka J. 27 Talandová M. 4 Teresa Sesma M. 29 Tesařová M. 40 Tolde Z. 41

Tremlová B. 4 Tyrpekl V. 7 Uhrík B. 10 Vacková T. 42 Vancová M. 21 Vávra I. 17, 36, 37, 43 Venclíková Z. 21 Venit T. 35 Vlková H. 39 Vystavěl T. 10 Wandrol P. 23 Watanabe K. 8 Weyda F. 4 Zárubová N. 8 Žďánský K. 2

79

Adresář účastníků

Anger Martin Ústav živočišné fyziologie a genetiky Rumburská 89 27721 Liběchov Tel: 315639591 e-mail: [email protected]

Černohorský Ondřej ČVUT FJFI Břehová 7 11519 Praha 1 Tel: 603 209 598 e-mail: [email protected]

Baumann Manfred MBSS Science Service Banská Hodruša 485 96663 Sloveksno Tel: 421456 84 40 83 e-mail: [email protected]

Chernyavskiy Oleksandr Fyziologický ústav AVČR v.v.i. Videnska 1083 14200 Praha 4 Tel: 241062274 e-mail: [email protected]

Benada Oldřich Mikrobiologický ústav AV ČR, v.v.i. Vídeňská 1083 142 20 Praha Tel: 241 06 23 99 e-mail: [email protected]

Čiampor Fedor Virologický ústav SAV Dúbravská cesta 9 845 05 Bratislava Tel: 00421 2 59302454 e-mail: [email protected]

Bílý Tomáš Parazitologický ústav AVČR vvi – LEM Branišovská 31 37005 České Budějovice Tel: 387 77 59 94 e-mail: [email protected]

Čiamporová Milada Botanický ústav SAV Dúbravská cesta 9 845 23 Bratislava Tel: 00421-2-5942 6129 e-mail: [email protected]

Brázda Václav Tel: 541517231 e-mail: [email protected]

Dammer Jiří ÚTEF ČVUT Horská 3a/22 128 00 Praha 2 Tel: 420224 35 91 79 e-mail: [email protected]

Brázdová Eva Tel: 541517257 e-mail: [email protected]

80

Dimzoski Bojan Ústav makromolekulární chemie Heyrovského nám. 2 162 06 Praha Tel: 296 80 92 68 e-mail: [email protected]

Gába Alexandr Specion s.r.o. Budějovická 55 14000 Praha 4 Tel: 602366406 e-mail: [email protected]

Dzijak Rastislav Ustav molekularni genetiky AV ČR Videnska 1083 14220 Praha Tel: 241 06 31 54 e-mail: [email protected]

Glatzová Hana TESCAN, s.r.o. Libušina třída 21 623 00 Brno Tel: 547 13 04 14 e-mail: [email protected]

Filimonenko Anatoly Ústav molekularní genetiky AV ČR Vídeňská 1083 142 20 Praha 4 Tel: 420241 06 31 52 e-mail: [email protected]

Gnauck Peter Carl Zeiss spol. s r.o. Radlická 14/3201 150 00 Praha 5 Tel: 420233 10 12 35 e-mail: [email protected]

Filimonenko Vlada Ústav molekularní genetiky AV ČR Vídeňská 1083 142 20 Praha 4 Tel: 420241 06 31 53 e-mail: [email protected]

Gramblička Michal Fakulta chem. a potr. technológie Radlinského 9 812 37 Bratislava Tel: +421 915 747 892 e-mail: [email protected]

Fischerová Jana Medzinárodné laserové centrum Ilkovičova 3 841 04 Bratislava Tel: 421908502331 e-mail: [email protected]

Grym Jan Ústav fotoniky a elektroniky, AVČR Chaberska 57 18251 Praha 8 Tel: 266 77 34 17 e-mail: [email protected]

Flodrová Eva UPT AV ČR, v.v.i Královopolská 147 61264 Brno Tel: 420541 51 43 47 e-mail: [email protected]

Havránek Vladimír Ústav jaderné fyziky AVČR v.v.i. 250 68 Řež u Prahy Tel: 266173127 e-mail: [email protected] Hlavenka Petr FEI Cezech Republic Podnikatelska 6 612 00 Brno Tel: 533 311 226 e-mail: [email protected]

Frank Luděk ÚPT AV ČR, v.v.i. Královopolská 147 61264 Brno Tel: 541 51 42 99 e-mail: [email protected] 81

Hodová Iveta Přírodovědecká fakulta, Masarykova univerzita Kotlářská 2 611 37 Brno Tel: 420549 49 46 64 e-mail: [email protected]

Hudeček František Carl Zeiss spol. s r.o. Radlická 14/3201 150 00 Praha 5 Tel: 420233 10 12 35 e-mail: [email protected] Jäger Aleš Fyzikální ústav AV ČR, v. v. i. Na Slovance 2 182 21 Praha Tel: 241062424 e-mail: [email protected]

Holčáková Regina VŠB-TU Ostrava, FEI, Katedra Elektroenergetiky 17. Listopadu 15 70200 Ostrava-Poruba Tel: 607 16 83 74 e-mail: [email protected]

Janáček Jiří Fyziologický ústav AVČR Vídeňská 1083 142 00 Praha 4 Tel: 241062424 e-mail: [email protected]

Horáček Miroslav Ústav přístrojové techniky AV ČR Královopolská 147 61264 Brno Tel: 420541 51 43 18 e-mail: [email protected]

Janda Pavel Ústav fyzikální chemie J. Heyrovského Dolejškova 3 182 23 Praha 8 Tel: 266053966 e-mail: [email protected]

Horák Radovan PE Systems, s.r.o. Pastevců 471 14900 Praha Tel: 420724 13 46 32 e-mail: [email protected]

Janoušek Karel ÚMG AV ČR, v. v. i. Vídeňská 1083 142 20 Praha 4 Tel: 420 241 06 31 61 e-mail: [email protected]

Hovorka Miloš ÚPT AV ČR, v.v.i Královopolská 147 61264 Brno 3 Tel: 541 514 259 e-mail: [email protected]

Jelínková Iva Ústav molekulární genetiky Vídeňská 1083 14220 Praha 4 Tel: 241 063 152 e-mail: [email protected]

Hozák Pavel Ústav molekulární genetiky AV ČR Vídeňská 1083 14220 Praha 4 Tel: 420603 87 28 72 e-mail: [email protected]

Klein Pavel UPT AV ČR Královopolská 147 51264 Brno Tel: 541 514 111 e-mail: [email protected]

Hromádková Jiřina Ústav makromolekulární chemie Heyrovského nám. 2 162 06 Praha Tel: 296 80 92 73 e-mail: [email protected] 82

Klementová Mariana UACH AVČR, v.v.i. Husinec-Řež 1001 250 68 Husinec-Řež Tel: 723526619 e-mail: [email protected]

Krist Pavel Carl Zeiss spol. s r.o. Radlická 14/3201 150 00 Praha 5 Tel: 233 101 235 e-mail: [email protected]

Klepetář Jan FGU AV ČR Vídenská 1083 142 20 Praha 4 Tel: +420 605 179 682 e-mail: [email protected]

Kříž Pavel ÚMG AV ČR Vídeňská 1083 142 20 Praha 4 Tel: 420296 44 31 54 e-mail: [email protected]

Kolařík Vladimír DELONG INSTRUMENTS a.s. Palackého třída 153b 61200 Brno Tel: 549 123 504 e-mail: [email protected]

Križanová Zuzana Elektrotechnický ústav SAV Dúbravská cesta 9 841 04 Bratislava Tel: +421 2 5922 2171 e-mail: [email protected]

Kolíbal Miroslav Fakulta strojního inženýrství Technická 2896/2 61669 Brno Tel: 541 14 28 13 e-mail: [email protected]

Ing. Vladislav Krzyžánek University of Muenster, Institute of Medical Physics and Biophysics Robert-Koch-Str. 31 D-48149 Muenster Tel: +49-251-83 55137 [email protected]

Konvalina Ivo Ústav přístrojové techniky AV ČR Královopolská 147 61264 Brno Tel: 420541 51 42 59 e-mail: [email protected]

Kubíčková Kristýna Tescan, a.s. Libušina třída 21 623 00 Brno Tel: 547 130 475 e-mail: [email protected]

Kopřiva Radomír Tescan, a.s. Libušina třída 21 623 00 Brno Tel: 547 130 470 e-mail: [email protected]

Kubínová Lucie Fyziologický ústav AV ČR Vídeňská 1083 14220 Praha 4 Tel: 241 06 23 14 e-mail: [email protected]

Kozubek Michal Masarykova univerzita Fakulta informatiky Botanická 68a 60200 Brno Tel: 420549494023 e-mail: [email protected]

83

Kubínová Zuzana Přírodovědecká fakulta Univerzity Karlovy Albertov 6 128 43 Praha 2 Tel: 221 951 694 e-mail: [email protected]

Martinka Michal Faculty of Natural Sciences, Comenius University in Bratislava Mlynská dolina B2 84215 Bratislava Tel: 421260 29 64 55 e-mail: [email protected]

Kunka Igor UMI ČVUT v Praze Technicka 4 16600 Praha 6 Tel: +420 725 888 650 e-mail: [email protected]

Mašová Šárka Přírodovědecká fakulta MU, ÚBZ Kotlářská 2 611 37 Brno Tel: 420549 49 44 47 e-mail: [email protected]

Langhans Jan Parazitologický ústav AV Branišovská 31 31 37005 České Budějovice Tel: 420387775995 e-mail: [email protected]

Matula Pavel Masarykova Univerzita Botanická 68a 60200 Brno e-mail: [email protected] Michalcová Alena VŠCHT Praha Technická 5 166 28 Praha 6 Tel: 420220443683 e-mail: [email protected]

Lednický František Ústav makromolekulární chemie AV ČR Heyrovského nám. 2 162 06 Praha 6 Tel: 296809357 e-mail: [email protected]

Michálek Jan FgÚ AV ČR Vídeňská 1083 14220 Praha Tel: +420 607 184 642 e-mail: [email protected]

Lukeš Jaroslav RMI, s. r. o. Perštýnská 116 53341 Lázně Bohdaneč Tel: 602261493 e-mail: [email protected]

Mika Filip ÚPT AV ČR Královopolská 147 61200 Brno Tel: 541514298 e-mail: [email protected]

Marášek Pavel Ústav molekulární genetiky AV ČR Vídeňská 1083 142 20 Praha 4 Tel:241 063 152 e-mail: [email protected]

Müllerová Ilona Ústav přístrojové techniky AVČR Královopolská 147 61264 Brno Tel: 541514300 e-mail: [email protected]

Marek Martin VSB-TU Ostrava, FEI, KAT410 17. listopadu 15 70800 Ostrava-Poruba Tel: 724 03 37 27 e-mail: [email protected] 84

Munzar Martin RMI s.r.o. Pernštýnská 116 53341 Lázně Bohdaneč Tel: 602472999 e-mail: [email protected]

Palček Peter KMI, SjF, Žilinská univerzita Univerzitná 8215/1 010 26 Žilina Tel: 421415136004 e-mail: [email protected]

Nebesářová Jana Biologické centrum AV ČR Branišovská 31 370 05 České Budějovice Tel: 387 77 54 02 e-mail: [email protected]

Pelc Radek Institute of Physiology v.v.i. Vídenská 1083 14200 Praha 4 Tel: 2-4106-3766 e-mail: [email protected]

Ing. Vilém Neděla UPT AV ČR Královopolská 147 61264 Brno Tel: 420541 51 43 33 e-mail: [email protected]

Peřina Vratislav Ústav jaderné fyziky AV ČR Řež 000 250 68 Řež u Prahy Tel: 266172106 e-mail: [email protected]

Nevoralová Martina Ústav makromolekulární chemie AV Heyrovského náměstí 2 162 06 Praha 6 Tel: 296 809 261 e-mail: [email protected]

Piksová Kateřina ČVUT v Praze Břehová 7 11519 Praha 1 Tel: 221912817 e-mail: [email protected]

Novák Ondřej UEM AV ČR Víděňská 1083 14220 Praha Tel: 777249535 e-mail: [email protected]

Pišlová Lenka Ústav molekulární genetiky AV ČR Vídeňská 1083 14220 Praha 4 – Krč Tel: 241 06 22 89 e-mail: [email protected]

Novotný Filip Fakulta jaderná a fyzikálně inženýrská Břehová 7 11519 Praha 1 Tel: 221912823 e-mail: [email protected]

Plášek Jaromír Univerzita Karlova v Praze, MFF Ke Karlovu 3 12116 Praha Tel: 723471126 e-mail: [email protected]

Ovečka Miroslav Botanický ústav SAV Dúbravská cesta 9 845 23 Bratislava Tel: +421 2 59426134 e-mail: [email protected]

Pokorná Zuzana Ústav přístrojové techniky AV ČR Královopolská 147 61264 Brno Tel: 420541 51 43 16 e-mail: [email protected]

85

Pospiech Matej VFU Brno Palackeho 1-3 61242 Brno Tel: 541562704 e-mail: [email protected]

Rusnák Andrej Ústav molekulárnej fyzioógie a genetiky SAV Vlárska 5 83334 Bratislava Tel: 421918409658 e-mail: [email protected]

Proška Jan Fakulta jaderná a fyzikálně inženýrská ČVUT v Praze Břehová 7 115 19 Praha 1 Tel: 221912416 e-mail: [email protected]

Říhová Zuzana ČVUT v Praze Technická 4 16607 Praha Tel: 604 7838 72 e-mail: [email protected]

Radochová Barbora Fyziologický ústav AV ČR Vídeňská 1083 142 20 Praha 4 - Krč Tel: 296443769 e-mail: [email protected]

Sedlák Ondřej Nikon, s.r.o. Kodaňská 46 10000 Praha Tel: 724556731 e-mail: [email protected]

Richter Mark SPECION s.r.o./ JPK Instruments AG Bouchéstr. 12 D-12435 Berlin Tel: +49 305 331 12070 e-mail: [email protected]

Semelbauer Marek Ústav zoológie Slovanská akadémia vied Dúbravská cesta 9 845 06 Bratislava Tel: 0914 257735 e-mail: [email protected]

Rohožková Jana Ústav molekulární genetiky AV ČR Vídeňslá 1083 14220 Praha Tel: 420241 06 31 53 e-mail: [email protected]

Schröfel Adam VŠB-TUO, Centrum nanotechnologií 17. listopadu 15 70833 Ostrava Tel: 420777864404 e-mail: [email protected]

Rous Zdeněk MIKRO, spol. s r.o. Lísky 94 624 00 Brno Tel: 603 48 25 09 e-mail: [email protected]

Sobol Margaryta Sobol Ústav molekulární genetiky AV ČR Vídeňská 1083 14220 Praha Tel: 420 241 06 31 52 e-mail: [email protected]

Rozkošný Ivan Nikon spol. s r.o. Kodaňská 46 100 00 Praha Tel: 602363767 e-mail: [email protected]

Sopko Vítek ÚTEF ČVUT Horská 3a/22 128 00 Praha Tel: 420 224 35 91 79 e-mail: [email protected] 86

Srbková Zuzana JEOL (EUROPE) SAS-organizační složka Karlovo náměstí 13 121 35 Praha 2 Tel: 420603221751 e-mail: [email protected]

Tesařová Martina Biologické centrum Branišovská 31 37005 České Budějovice Tel: 604690897 e-mail: [email protected]

Svoboda David Masarykova univerzita Botanická 68a 628 00 Brno e-mail: [email protected]

Tolde Zdeněk ČVUT v Praze Technická 4 16607 Praha 6 Tel: 777819782 e-mail: [email protected]

Sýkora Jiří UPT AV ČR Brno Královopolská 147 61264 BRNO Tel: +420 541 514 111 e-mail: [email protected]

Tyrpekl Václav UACH AV CR,v.v.i Husinec-Rez 1001 25068 Rez Tel: 266173144 e-mail: [email protected]

Šebesta Ondřej Přírodovědecká fakulta Albertov 6 12843 Praha Tel: 221951943 e-mail: [email protected]

Unčovský Marek FEI Czech Republic s.r.o. Podnikatelská 6 612 00 Brno Tel: 420533311236 e-mail: [email protected]

Šittner Petr Fyzikální ústav AV ČR Na Slovance 2 18221 Praha Tel: 286 890 527 e-mail: [email protected]

Vacková Taťana Ústav makromolekulární chemie Heyrovského nám. 2 162 06 Praha 6 Tel: 222512161 e-mail: [email protected]

Šlouf Miroslav Ústav makromolekulární chemie Heyrovského nám. 2 162 06 Praha Tel: 296 80 92 91 e-mail: [email protected]

Vávra Ivo Elektrotechnický ústav SAV Dúbravská cesta 9 841 04 Bratislava Tel: +421 2 5922 2889 e-mail: [email protected]

Švec Peter Fyzikálny Ústav SAV Dubravská cesta 9 84511 Bratislava Tel: +421 2 59410561 e-mail: [email protected]

Venit Tomáš Ústav molekulární genetiky Vídeňská 1083 14220 Praha 4 Tel: 420241 06 31 54 e-mail: [email protected]

87

Vlková Helena Ústav makromolekulární chemie Heyrovského nám. 2 162 06 Praha Tel: 296 80 92 68 e-mail: [email protected]

Weyda František Biologické centrum AV ČR, v.v.i. Branišovská 1160/31 37005 České Budějovice Tel: 606-651179 e-mail: [email protected]

Vystavěl Tomáš FEI Podnikatelská 6 61200 Brno Tel: 420 737 20 80 91 e-mail: [email protected]

Zárubová Niva Fyzikální ústav AVČR v.v,i. Na Slovance 1999/2 182 21 Praha 8 Tel: 266 052 613 e-mail: [email protected]

Wandrol Petr FEI Podnikatelská 6 612 00 Brno Tel: 739003189 e-mail: [email protected]

Zemek Alexandr EDLIN, s.r.o. Za Kralupkou 440 27711 Libiš Tel: 267 10 82 55 e-mail: [email protected]

88

Adresář vystavovatelů

Carl Zeiss spol. s.r.o. Radlická 14/3201 150 00 Praha 5 Smíchov Tel.: (+420) 233 101 221 e-mail: [email protected]

Nikon s.r.o. Kodaňská 46 100 10 Praha 10 Tel: 224 315 650 e-mail: [email protected]

Edlin s.r.o. Za Kralupkou 440 277 11 Libiš Tel.: 267 108 255 e-mail: [email protected]

PE Systems s.r.o. Pastevců 471 149 00 Praha 4 Tel: 241 430 534 e-mail: [email protected]

FEI Czech Republic s.r.o. Podnikatelská 6 612 00 Brno Tel.: +420 541 192 222 e-mail: [email protected]

RMI s.r.o. Pernštýnská 116 533 41 Lázně Bohdaneč Tel: 466 921 885 e-mail: [email protected]

JEOL (EUROPE) s.r.o. Karlovo náměstí 13 121 35 Praha 2 Tel: 224 916 714 e-mail: [email protected]

Specion s.r.o. Budějovická 55 140 00 Praha 4 Tel: 244 402 091 e-mail: [email protected]

Mikro s.r.o. Dolnokrčská 54 140 00 Praha 4 – Krč Tel: 267 108 255 e-mail: [email protected]

Tescan a.s. Libušina tř. 21 623 00 Brno Tel: +420 547 130 411 e-mail: [email protected]

89

Poznámky

90