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Research Institute of Biomolecule Metrology Co., Ltd.

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High-Speed Atomic Force Microscope - Ando model -




NanoExplorer (NEX)


Dynamic Visualization of nano-scale world

HS-AFM*1.0 - Ando model - is the High-Speed Atomic Force Microscope which was developed based on the research achievements accomplished by Prof. Ando in Kanazawa University. This is the world’s first instrument that broke through the weak point of conventional AFM “low-speed”, and realized the video rate scan. The high-speed scan enables us to capture swinging molecules in solution clearly without blurring. Consequently, the strong anchoring of a sample to the substrate is unnecessary and a dynamic observation is achieved without losing the activities of soft biomolecules.


*The HS-AFM was developed by Prof. Ando (Kanazawa Univ.) and commercialized by RIBM.


Walking_myosinV
Walking_myosinV
bacteriorhodopsin (D96N)
rotorless F1-ATPase
walking myosin V(realtime)
bacteriorhodopsin
in response to light(x10)
rotorless F1-ATPase(realtime)

Contents


High-speed scanner

  • Active dumping and counter balance can suppress mechanical vibration.
  • Compact and reliable design

In addition to Standard high-speed scanner, specialized scanners:

Injection type, Ultra high-speed, Wide scanner are optionally available.

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Ultra small and soft cantilever with high resonance frequency

  • Cantilever for HS-AFM can realize the observation of any soft samples such as a protein.
 

Ultra small cantilever

Resonance frequency :1500 kHz (in air )

Force constant :0.1 N/m

Tip radius :< 10 nm

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High-speed and stable feedback control

  • Clear and fine images of surface structures can be obtained by using the high-speed scanning.

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Movie Gallery

IgG

IgG antibodies


Dynamic Imaging of the IgG antibodies
DNA

Plasmid DNA


Dynamic Imaging of the Plasmid DNA on the mica-substrate
DNase

Dynamic Imaging of the DNA digestion process by DNase


(DNA : λDNA , DNase: endonuclease)
DNA digestion by Nuclease

DNA digestion by Nuclease


Dynamic Imaging of the DNA digestion process
(DNA : pUC18 plasmid DNA , DNA digestion: Bal31 nuclease)
DNA Polymerase reaction

DNA Polymerase reaction


Dynamic Imaging of the DNA elongation process with Phi29 in liquid
(DNA:λDNA , DNA polymerase: Phi29)
Point defect in streptavidin 2D crystal

Point defect in streptavidin 2D crystal


Dynamic Imaging of the diffusion on point defects in the crystal

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Image Gallery

IgG
plasmid DNA
myosinⅡ
IgG antibody
150nm * 150nm
Standard high-speed scanner
plasmid DNA
250nm * 250nm
Standard high-speed scanner
myosinⅡ
500nm * 500nm
Standard high-speed scanner
streptavidin
GroEL
bacteriorhodopsin
streptavidin
90nm * 90nm
(*1)
GroEL
90nm * 90nm
(*1)
bacteriorhodopsin
40nm * 40nm
(*1)
lipid membrane
350nm Beads
350nm Beads
lipid membrane
3500nm * 3500nm
Wide scanner
350nm polystyrene beads
3000nm * 3000nm
Wide scanner
350nm polystyrene beads
900nm * 900nm
Wide scanner
ecoli
E.coli
3000nm * 3000nm
Wide scanner

(*1) Image courtesy of Prof. Ando (Kanazawa Univ.)


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Published Papers

Video imaging of walking myosin V




a. and b. : Direct observation of walking myosin V. a. 130 nm x 65 nm, b. 125 nm x 62.5 nm. c. Scheme of myosin V walking.


Myosin V is a two-headed processive motor and functions as cargo transporter in cells. The dynamic behaviors of myosin V translocating along actin filaments were visualized by HS-AFM. The high-resolution movies provided not only ‘visual evidence’ for previously speculated or demonstrated molecular behaviors, including lever-arm swing, but also more detailed behaviors of the molecules, leading to a comprehensive understanding of the motor mechanism. This direct and dynamic high-resolution visualization is a powerful new approach to studying the structure and dynamics of biomolecules in action.

N. Kodera et al. Nature 468, 72 (2010). Kanazawa University


Dynamic molecular processes in photoactivated bacteriorhodopsin



Structural change of bacteriorhodopsin (bR), which is known as light-driven proton pump, has been visualized by HS-AFM. The purple membrane composed of D96N bR mutant, which has a longer photocycle (10’s) than that of the wild type, was adsorbed on mica substrate. Upon illumination with green light (532nm), bR drastically changes its structure and returns to the unphotolysed state in a few seconds after light-off. This outcome is reproducible in repeated dark illumination cycles.

M. Shibata et al. Nature Nanotech. 5, 208 (2010). Kanazawa University


Point defect in streptavidin 2D crystal




a. Dynamic observation of the diffusion of point defects. b. Scheme of streptavidin arrays in a C222 crystal. Unit lattice vectors are indicated by red arrows.


The diffusion of point defects in the crystals was successfully observed by HS-AFM. In Figure a, the trajectory tracking of two monovacancy defects was obviously anisotropic with respect to the two axes of the crystalline lattice. As a result, the diffusion constant D along each axis could be established to be Da = 20.5 nm2/s and Db = 48.8 nm2/s. This means, HS-AFM is useful for studying various dynamic process, such as the crystal growth and disintegration processes.

D. Yamamoto et al. Nanotechnology 19, 384009 (2008). Kanazawa University


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Reference List

Biological sample

The authors Title Journal
I. Casuso, N. Kodera, C. Le Grimellec, T. Ando and S. Scheuring Contact-mode high-resolution high-speed atomic force microscopy movies of the purple membrane Biophys., J. 97, 1354 (2009).
J. L. Gilmore, Y. Suzuki, G. Tamulaitis, V. Siksnys, K. Takeyasu and Y. L. Lyubchenko Single-molecule dynamics of the DNA-EcoRII protein complexes revealed with high-speed atomic force microscopy Biochemistry, 48, 10492 (2009).
M.-C. Giocondi, D. Yamamoto, E. Lesniewska, P.-E. Milhiet, T. Ando, and C. L. Grimellec Surface topography of membrane domains BBA-Biomembranes, 1978, 703 (2010).
Y. L. Lyubchenko, L. S. Shlyakhtenko and A. A. Gall Atomic force microscopy imaging and probing of DNA, proteins, and protein DNA complexes: silatrane surface chemistry Methods mol. Biol., 543, 337 (2009).
Y. L. Lyubchenko, L. S. Shlyakhtenko AFM for analysis of structure and dynamics of DNA and protein-DNA complexes Methods mol. Biol., €47, 206 (2009)
L. S. Shlyakhtenko, A. Y. Lushnikov, Y. L. Lyubchenko Dynamics of Nucleosomes Revealed by Time-Lapse Atomic Force Microscopy Biochemistry,€48, 7842 (2009)
N. Kodera, D. Yamamoto, R. Ishikawa and T. Ando Video imaging of walking myosin V by high-speed atomic force microscopy Nature, 468, 72 (2010).
S. Sugimoto, K. Yamanaka, S. Nishikori, A. Miyagi, T. Ando and T. Ogura AAA Chaperone ClpX Regulates Dynamics of Prokaryotic Cytoskeletal Protein FtsZ J. Biol. Chem., 285, 6648 (2010).
D. Yamamoto, N. Nagura, S. Omote, M. Taniguchi and T. Ando Streptavidin 2D crystal substrates for visualizing biomolecular processes by atomic force microscopy Biophys. J., 97, 2358 (2009).
M. Shibata, H. Yamashita, T. Uchihashi, H. Kandori and T. Ando High-speed atomic force microscopy shows dynamic molecular processes in photoactivated bacteriorhodopsin Nature Nanotech., 5, 208 (2010).
D. Yamamoto, T. Uchihashi, N. Kodera, H. Yamashita, S. Nishikori, T. Ogura, M. Shibata and T. Ando High-speed atomic force microscopy techniques for observing dynamic biomolecular processes Meth. Enzymol., 475, 541 (2010).
P.-E. Milhiet, D. Yamamoto, O. Berthoumieu, P. Dosset, C. L. Grimellec, J.-M. Verdier, S. Marchal, and T. Ando Deciphering the structure, growth and assembly of amyloid-like fibrils using high-speed atomic force microscopy PLos One, 5, e13240 (2010).
H. Yamashita, K. Voitchovsky, T. Uchihashi, S. A. Contera, J. F. Ryan , Ando T. Dynamics of bacteriorhodopsin 2D crystal observed by high-speed atomic force microscopy J. Struct. Biol., 167, 153 (2009).
D. Yamamoto, T. Uchihashi, N. Kodera and T. Ando Anisotropic diffusion of point defects in a two-dimensional crystal of streptavidin observed by high-speed atomic force microscopy Nanotechnology, 19, 384009 (2008).
T. Ando, T. Uchihashi, N. Kodera, D. Yamamoto, A. Miyagi, M. Taniguchi and H. Yamashita High-speed AFM and nano-visualization of biomolecular processes Eur. J. Physiol., 456, 211 (2008).
A. Miyagi, Y. Tsunaka, T. Uchihashi, K. Mayanagi, S. Hirose, K. Morikawa, and T. Ando Visualization of Intrinsically Disordered Regions of Proteins by High-Speed Atomic Force Microscopy Chemphyschem., 9, 1859 (2008).
K. Shinohara, N. Kodera and T. Ando Single Molecular Imaging of a micro-Brownian Motion and a Bond Scission of a Supramolecular Chiral π-Conjugated Polymer as a Molecular Bearing Driven by Thermal Fluctuations Chem. Lett., 36, 1378 (2007).
H. Yamashita, N. Kodera, A. Miyagi, T. Uchihashi, D. Yamamoto and T. Ando Tip-sample distance control using photothermal actuation of a small cantilever for high-speed atomic force microscopy Rev. Sci. Instrum. 78, 083702 (2007).
T. Ando, T. Uchihashi, N. Kodera, A. Miyagi, R. Nakakita, H. Yamashita and M. Sakashita High-Speed Atomic Force Microscopy for Studying the Dynamic Behavior of Protein Molecules at Work J. J. Appl. Phys., 45, 1897 (2006).
S. Morita, H. Yamada and T. Ando Japan AFM roadmap 2006 Nanotechnology, 18, 084001 (2007).
H. Koide, T. Kinoshita, Y. Tanaka, S. Tanaka, N. Nagura, G. Meyer zu Ho¨rste, A. Miyagi and T. Ando Identification of the Single Specific IQ Motif of Myosin V from Which Calmodulin Dissociates in the Presence of Ca2+ Biochemistry, 26, 11598 (2006).
Mikihiro Shibata, Takayuki Uchihashi, Hayato Yamashita, Hideki Kandori, and Toshio Ando Structural Changes in Bacteriorhodopsin in Response to Alternate Ilumination Observed by High-Speed Atomic Force Microscopy Angew. Chem. Int. Ed. 50, 4410-4413 (2011).
Takayuki Uchihashi, Ryota Iino, Toshio Ando Hiroyuki Noji High-Speed Atomic Force Microscopy Reveals Rotary Catalysis of Rotorless F1-ATPase Science 333, 1279 (2011).
Y. Shinozaki, A. M. Siitonen, K. Sumitomo, K. Furukawa, and K. Torimitsu Effect of Ca2+ on Vesicle Fusion on Solid Surface: An In vitro Model of Protein-Accelerated Vesicle Fusion Jpn. J. Appl. Phys., 47, 6164 (2008).
Y. Shinozaki, K. Sumitomo, M. Tsuda, S. Koizumi, K. Inoue, K. Torimitsu Direct Observation of ATP-Induced Conformational Changes in Single P2X4 Receptors Plos Biol. 7, e1000103 (2009).
Y. Shinozaki, K. Sumitomo, K. Furukawa, H. Miyashita, Y. Tamba, N. Kasai, H. Nakashima and K. Torimitsu Visualization of Single Membrane Protein Structure in Stretched Lipid Bilayer Suspended over Nanowells Appl. Phys. Express. 3. 027002 (2010)
H. Sugasawa, Y. Sugiyama, T. Morii and T. Okada Dynamic Observation of 2686bp DNA-BAL 31 Nuclease Interaction with Single Molecule Level Using High-Speed Atomic Force Microscopy Jpn. J. Appl. Phys., 47, 6168 (2008).
S.-I. Yamamoto, T. Okada, Y. Uraoka, I. Yamashita and S. Hasegawa Static and dynamic observation of supermolecular protein, ferritin, using high-speed atomic force microscope. J. Appl. Phys., 109, 034901 (2011)
Y. Suzuki, Y. Higuchi, K. Hizume, M. Yokokawa, S. H. Yoshimura, K. Yoshikawa and K. Takeyasu Molecular dynamics of DNA and nucleosomes in solution studied by fast-scanning atomic force microscopy Ultramicrosc., 110, 682 (2010)
M. Yokokawa, C. Wada, T. Ando, N. Sakai, A. Yagi, S. H. Yoshimura and K. Takeyasu Fast-scanning atomic force microscopy reveals the ATP/ADP-dependent conformational changes of GroEL EMBO J., 25, 4567 (2006).
N. Crampton, M. Yokokawa, D. T. F. Dryden, J. M. Edwardson, D. N. Rao, K. Takeyasu, S. H. Yoshimura, and R. M. Henderson Fast-scan atomic force microscopy reveals that the type III restriction enzyme EcoP15I is capable of DNA translocation and looping PNAS, 104, 12755 (2007).
F. Tanaka, T. Mochizuki, X. Liang, H. Asanuma, S. Tanaka, K. Suzuki, S. Kitamura, A. Nishikawa, K. Ui-Tei and M. Hagiya Robust and photocontrollable DNA capsules using azobenzenes Nano Lett., 10, 3560 (2010).
K. Igarashi, A. Koivula, M. Wada, S. Kimura, M. Penttila and M. Samejima High speed atomic force microscopy visualizes processive movement of Trichoderma reesei cellobiohydrolase I on crystalline cellulose J. Biol. Chem., 284, 36186 (2009).
Kiyohiko Igarashi, Takayuki Uchihashi, Anu Koivula, Masahisa Wada, Satoshi Kimura,Tetsuaki Okamoto, Merja Penttila, Toshio Ando, Masahiro Samejima1 Traffic Jams Reduce Hydrolytic Efficiency of Cellulase on Cellulose Surface Science 333, 755 (2011)
Toshio Ando Observation of the protein molecule by HS-AFM Applied physics, 77, 1181 (2008).
M. Endo and H. Sugiyama Three-dimensional DNA nanostructures constructed by folding of multiple rectangles Nucleic Acids Symposium Series, 53, 81 (2009).
M. Endo, Y. Katsuda, K. Hidaka and H. Sugiyama Regulation of DNA Methylation Using Different Tensions of Double Strands Constructed in a Defined DNA Nanostructure J. Am. Chem. Soc., 132, 1592 (2010).
M. Endo, T Sugita, Y. Katsuda, K. Hidaka and H. Sugiyama Programmed-assembly system using DNA jigsaw pieces Chem.-Eur. J., 16, 5362 (2010).
M. Endo and H. Sugiyama Chemical Approaches to DNA Nanotechnology. ChemBioChem, 10, 2420 (2009).
M. Endo, T. Sugita, A. Rajendran, Y. Katsuda, T. Emura, K. Hidaka and H. Sugiyama Two-dimensional DNA origami assemblies using a four-way connector. Chem. Commun., 47, 3213 (2011)
Y. Sannohe, M. Endo, Y. Katsuda, K. Hidaka and H. Sugiyama Visualization of Dynamic Conformational Switching of the G-Quadruplex in a DNA Nanostructure J. Am. Chem. Soc., 132. 16311 (2010)
M. Endo, K. Hidaka and H. Sugiyama Direct AFM observation of an opening event of a DNA cuboid constructed via a prism structure Org. Biomol. Chem., DOI: 10.1039/ c0ob01093f (2011)
A. Rajendran, M. Endo, Y. Katsuda, K. Hidaka and H. Sugiyama Programmed Two-Dimensional Self-Assembly of Multiple DNA Origami Jigsaw Pieces ACS Nano, 5, 665 (2011)
Wickham SF, Endo M, Katsuda Y, Hidaka K, Bath J, Sugiyama H, Turberfield AJ Direct observation of stepwise movement of a synthetic molecular transporter. Nat Nanotechnol. 2011 Feb 6.

Other

The authors Title Journal
T. Itani and J. J. Santillan In situ Characterization of Photoresist Dissolution Appl. Phys. Exp., 3, 061601 (2010).
T. Itani and J. J. Santillan Dissolution Behavior of Photoresists: An In-situ Analysis J. Photopoly. Sci. Technol., 23, 639 (2010).
Shigeto Inoue,a Takayuki Uchihashi, Daisuke Yamamotob and Toshio Ando Direct observation of surfactant aggregate behavior on a mica surface using high-speed atomic force microscopy Chem. Commun., 47, 4974 4976 (2011)
KEN-ICHI SHINOHARA, NORIYUKI KODERA, TAKASHI OOHASHI Single-Molecule Imaging of Photodegradation Reaction in a Chiral Helical π-Conjugated Polymer Chain Polymer Chemistry, 48, 4103 4107 (2010)

AFM development

The authors Title Journal
T. Fukuma, Y. Okazaki, N. Kodera, T. Uchihashi and T. Ando High resonance frequency force microscope scanner using inertia balance support Appl. Phys. Lett., 92, 243119 (2008)
T. Ando, T. Uchihashi and T. Fukuma High-speed atomic force microscopy for nano-visualization of dynamic biomolecular processes Prog. Surf. Sci., 83, 337 (2008).
T. Ando, T. Uchihashi1, N. Kodera, D. Yamamoto, M. Taniguchi, A. Miyagi1 and H. Yamashita High-speed atomic force microscopy for observing dynamic biomolecular processes J. Mol. Recognit., 20, 448 (2007).
T. Uchihashi, N. Kodera, H. Itoh, H. Yamashita and T. ANDO Feed-Forward Compensation for High-Speed Atomic Force Microscopy Imaging of Biomolecules J. J. Appl. Phys. 45, 1904 (2006).
N. Kodera, M. Sakashita and T. Ando Dynamic proportional-integral-differential controller for high-speed atomic force microscopy Rev. Sci. Instrum., 77, 083704 (2006).
N. Kodera, H. Yamashita and T. Ando Active damping of the scanner for high-speed atomic force microscopy Rev. Sci. Instrum., 76, 053708 (2005).
T. Ando, N. Kodera, T. Uchihashi, A. Miyagi, R. Nakakita, H. Yamashita and K. Matada High-speed Atomic Force Microscopy for Capturing Dynamic Behavior of Protein Molecules at Work J. Surf. Sci. Nanotech., 3 384 (2005).
T. Ando, N. Kodera, Y. Naito, T. Kinoshita, K. Furuta and Y. Y. Toyoshima A High-speed Atomic Force Microscope for Studying Biological Macromolecules in Action Chemphyschem., 4, 1196 (2003).
N. Kodera, T. Kinoshita, T. Ito and T. Ando High-resolution imaging of myosin motor in action by a high-speed atomic force microscope Adv. Exp. Med. Biol., 538, 119 (2003).
T. Ando, N. Kodera, D. Maruyama, E. Takai, K. Saito and A. Toda A High-Speed Atomic Force Microscope for Studying Biological Macromolecules in Action Jpn. J. Appl. Phys., 41, 4851 (2002).
T. Ando, N. Kodera, E. Takai, D. Maruyama, K. Saito and A. Toda A high-speed atomic force microscope for studying biological macromolecules PNAS, 98, 12468 (2001).
Toshio Ando, Noriyuki Kodera High speed video rate, AFM Measurement and control, 45, 2

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Standard system Specifications

Scan Speed 80 ms / frame (12.5 frames / sec)
Piezo Range X: 0.7 μm, Y: 0.7 μm, Z: 0.4 μm
Sample Size 1.5 mm in diameter
Detection Method Optical lever method
Scanning Method Sample scan
Environment In liquid
Control System PID control, Dynamic PID control
Measurement Mode AC mode. Topography and phase image
Significant Function Scanner active dumping
Drift correction for cantilever excitation

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Options

  • Units
Light irradiation Unit Light irradiation unit for the experiments with caged compounds.
Variable wavelength : 350 - 560 nm
Circulation unit The observation solutions can be exchanged while continuing AFM observation. The pH value and/or the buffer composition can be gradually changed during the measurement.
Heat control unit From room temperature to 50℃
*Under developments

  • Scanners
Ultra high-speed scanner Suitable for observing reactions between molecules .
  • Scan Speed : 50 ms / frame (20 frames / sec)
  • Scan Range : XY : 0.6 μm × 0.6 μm, Z : 0.2 μm
Wide scanner Suitable for relatively large samples with a high scanning rate.
  • Scan Speed : 1 s / frame (1 frame / sec)
  • Scan Range : XY : 4 μm × 4 μm, Z : 0.7 μm
Mechanically amplified
ultra wider scanner
Suitable for observing whole body of large samples such as cells.
  • Scan Speed : 10 s / frame (0.1 frames / sec)
  • Scan Range : XY : 30 μm × 30 μm, Z : 1.2 μm
* Each scan range is typical value.
** Maximum scanning rate is for a specific measurement condition.

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Contact

Research Institute of Biomolecule Metrology Co., Ltd.

807-133 Enokido, Tsukuba, Ibaraki 305-0853, Japan

Tel: +81-29-839-4611 Fax: +81-29-839-4612

E-mail: world-sales@ribm.co.jp


System configuration

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 Research Institute of Biomolecule Metrology Co., Ltd. is a trading, R&D and manufacturing-type venture capital organization. Our aim is to lead the technological advancement for measurements in nano-biology, a field which integrates biotechnology and nanotechnology. Through our bio-molecular measurement division, we develop various scanning probe microscopes that have unique features and reliable functionalities. With each device, our skilled staff can also provide operation software that is specialized to the customer’s need.

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