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Nanotechnology

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Nanotechnology Development and use of materials and devices for analysis and measurement on a nanometer scale Diverse and interdisciplinary field : – PowerPoint PPT presentation

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Title: Nanotechnology


1
Nanotechnology
  • Development and use of materials and devices
  • for analysis and measurement on a nanometer
    scale
  • Diverse and interdisciplinary field
  • Biology/Biotechnology, Physics, Chemistry,
    Material sciences, Electronics, Chemical
    Engineering, Information technology
  • Future implications
  • - Creation of new materials and devices in
    medicine, electronics,
  • energy etc.
  • - Social issues toxicity, environmental
    impact of nano-materials

2
Nanotechnology Plays by Different Rules
Normal scale
Nanoscale
3
Nano-sized materials
  • Unusual and different property
  • - Semiconductor nanocrystals
  • Size-dependent optical property Absorption
    and emission
  • - Magnetic nanoparticles (iron oxide)
  • Ferromagnetic Materials that can be
    magnetized by an
  • external magnetic filed and remain
    magnetized after the
  • external field is removed
  • The spin of the electrons in atoms
    is the main source of
  • ferromagnetism
  • Super-paramagnetic In the absence of
    magnetic field
  • ? Magnetization is
    zero
  • An external magnetic
    field is able to magnetize
  • the nanoparticles
  • ex) MRI contrast agents

4
Nano-Bio Convergence
Bio-inspired device and system
Bio-Technology
Nano-Technol
Molecular Imaging
Molecular Switch
DNA barcode
Biochip / Biosensor
Nanotherapy / Delivery
Bionano-machine / Nano-Robot
5
Nano-Biotechnology
  • Integration of nano-sized/structured materials,
    nano-scale analytical tools, and nano-devices
    into biological sciences including biotechnolgy
    for development of new biomaterials and
    analytical toolkits
  • Use of bio-inspired molecules or materials
  • - Typical characteristics of Biological
    events
  • - Self assembly
  • - Highly efficient high energy
    yield
  • - Very specific extremely
    precise
  • Bio-molecules
  • DNA, Proteins, Antibody, RNA, Aptamers,,
    Peptides,

6
Applications and perspectives of nanobiotechnology
  • Development of tools and methods
  • - More sensitive
  • - More specific
  • - Multiplexed
  • - More efficient and economic
  • Implementation
  • - Diagnosis and treatment of diseases
    Nanomedicine
  • Rapid and sensitive detection(Disease
    biomarkers, Imaging),
  • Targeted delivery of therapeutics
  • - Drug discovery
  • - Understanding of life science

7
Examples
  • Nano-Biodevices
  • Nano-Biosensors
  • Biomolecular motors
  • Medical use Targeting with Ab-magnetic
  • beads
  • Contrast agents for MRI
  • Analysis of a single molecule/ a single cell

8
Issues to be considered
  • Synthesis or selection of nano-sized/ structured
    materials Biocompatibility, cytotoxicity
  • Functionalization with biomolecules or for
    biocompatibility
  • Integration with devices and/or analytical tools
  • Assessment Reproducibility, Toxicity
  • Implementation to human body

9
The size of Things
10
How are nano-sized materials fabricated ?
11
NanoBiotech was initiated by development of AFM
and STM that enable imaging at atomic level in
1980
12
Atomic Force Microscope
Foremost tool for imaging, measuring, and
manipulating matter at the nanoscale
- When the tip is brought into proximity of a
sample surface, forces between the tip and the
sample lead to a deflection of the cantilever
according to Hooke's law
- Deflection is measured using a laser spot
reflected from the top surface of the cantilever
into an array of photodiodes
  • - If the tip was scanned at a constant height, a
    risk would exist that the tip collides with the
    surface, causing damage.
  • A feedback mechanism is employed to adjust the
    tip-to-sample distance to maintain a constant
    force between the tip and the sample.
  • Sample is mounted on a piezoelectric tube that
    can move the sample in the z direction for
    maintaining a constant force

- Contacting mode - Non-contacting mode - Tapping
mode
13
VEECO TESPA VEECO TESPA-HAR NANOWORLD SuperSharpSilicon

Tip length 10 ?m Radius 1520 nm Tip length 10 ?m (last 2 ?m 71) Radius 410 nm Tip length 10 ?m Radius 2 nm
14
Example showing the resolution of protein
structure by AFM
15
Optical Properties Of Quantum Dots
a) Multiple colors
b) Photostability
c) Wide absorption and narrow emission
d) High quantum yield
Quantum Yield 60 70
Single source excitation
16
In Vivo Cell Imaging

QD-Antibody conjugates
Antigen
? 3T3 cell nucleus stained with red QDs and
microtubules with green QDs
- Multiple Color Imaging - Stronger Signals
Wu et al. Nature Biotech. 2003 21 41
17
In Vivo Cell Imaging
Live Cell Imaging
Quantum Dot Injection
? Red Quantum Dot locating a tumor in a live mouse
Cell Motility Imaging
10um
? Green QD filled vesicles move toward to nucleus
(yellow arrow) in breast tumor cell
Alivisatos et al., Adv. Mater., 2002 14 882
18
Förster (or Fluorescence) Resonance Energy
Transfer (FRET)
  • Non-radiative energy transfer from an energy
    donor to an energy acceptor
  • Dipole dipole coupling
  • Energy transfer efficiency
  • - Degree of spectral overlap between donor
    fluorescence emission and acceptor
  • absorption
  • - 10 nm

19
Use of FRET measurements
Molecular ruler in the determination of inter- or
intra-molecular distances
  • Detection of target analytes
  • Analysis of biomolecular interactions
  • Single molecule analysis
  • - Protein folding/unfolding
  • - Protein dynamics
  • - The transfer efficiency is dependent on the
    inter-fluorophore distance.
  • Calculation of the transfer efficiency allows the
    distance between fluorophores.

20
FRET probe based on Quantum dots and AuNPs
  • QDs as an energy donor
  • - High quantum yield (0.6) and less
    photo-bleaching ? Strong PL intensity
  • - Multiple acceptors per single QD ?
    Increased ET efficiency
  • - Narrow and size-tunable emission ?
    Multiplexed assay
  • AuNPs as an energy acceptor (Quencher)
  • - Higher quenching efficiency than
    conventional quenchers
  • - Extended working distance by SET
    mechanism 22 nm
  • - Applicable to various donors

Oh et al., JACS (2006) Oh et al., Angewandte
Chemi Intl Ed.(2007)
21
Förster Resonance Energy Transfer (FRET) using QDs
Excitation
Excitation
Emission
FRET quenching
QD
QD
quencher
QD
quencher
QD
QD with Quencher
PL intensity
Time (ns)
22
Protease Assay
  • Protease
  • A variety of proteases in cells
  • Essential for the dynamic regulations of cell
    function and aberrations
  • Involved in major human diseases (cancers,
    apoptosis, and inflammation)
  • Protease inhibitors are known to be drug
    candidates
  • Matrix metalloproteinases (MMPs), Caspase-3,
    Thrombin
  • Conventional Methods for Protease Assay
  • Gel- and LC-MS-based methods accurate, but need
    a long analysis time
  • Limitation to high throughput assay
  • ? Simple, fast, and high throughput method is
    required

Assay of proteases using FRET probe -
Simple and sensitive - Multiplexed and
high-throughput assay
23
Matrix metalloproteinases (MMPs)
  • Regulate cancer metastasis by degrading ECM
    components or cytokines
  • MMP-7 Potential biomarker of human colorectal
    or breast cancer
  • MMP-2 9 Human carcinoma
  • MMP-3 Joint damage
  • MMP inhibitors Candidates for anticancer
    therapeutics

Steps in the process of metastasis
Nat. Rev. Cancer 2002, 2, 161
24
Construction of FRET probe for Protease Assay
Peptide-conjugated AuNP
FRET probe based on QDs and AuNPs
Kim et al., Anal. Chem. (2009)
25
Chip-based assay
Kim et al., Anal. Chem. (2009)
26
Chip-based Protease Assay
Fluorescence
Silver-staining
Relative PL ()
SA-QD605
(A)
SA-QD605 Pep-AuNPs
(B)
SA-QD605 Pep-AuNPs MMP-7 protease
(C)
SA-QD605 Pep-AuNPs MMP-7 protease Inhibitor
(D)
27
Specificity of FRET probe
Thrombin
MMP-7
Caspase-3
Peptide sequences for proteases Thrombin
Biotin-GKGGLVPR-GSGC MMP-7
Biotin-KSRWLA-LPRC Casp-3
Biotin-GRRGDEVD-GGGRRC
28
a -Hemolysin Self-Assembling Transmembrane Pore
  • A self-assembling bacterial exo-toxin produced by
    some pathogens like Staphylococcus aureus as a
    way to obtain nutrients ? lyzes red blood cells
  • Alpha-hemolysin monomers bind to the outer
    membrane of susceptible cells.
  • The monomers oligomerize to form a water-filled
    heptameric transmembrane channel that facilitates
    uncontrolled permeation of water, ions, and small
    organic molecules.
  • Rapid discharge of vital molecules, such as ATP,
    dissipation of the membrane potential and ionic
    gradients, and irreversible osmotic swelling
    leading to the cell wall rupture (lysis), can
    cause death of the host cell.

29
  • - Mushroom-like shape with a cap and stem
  • 50 A bets-barrel stem
  • Narrowest part 1.4 nm in diameter

30
Biotechnological applications
  • Unique structural features make self-assembling
    membrane channel
  • suitable for biotechnological applications
  • Assembled alpha-hemolysin stable over a wide
    range of pH and
  • temperature
  • Transmembrane pore stays open at normal
    conditions
  • Hemolysin binds to various biological or
    synthetic lipid bilayer
  • Delivery system facilitate controlled delivery
    of ions and small
  • organic compounds such as sugars and
    oligonucleotides across
  • the plasma membrane of cells or through the
    walls of synthetic
  • lipid vesicles Engineered pores
  • Stochastic sensors
  • DNA sequencing

31
Stochastic sensor
  • A molecular adaptor is placed inside its
    engineered stem,
  • influencing the transmembrane ionic current
    induced by an applied voltage
  • Reversible binding of analytes to the molecular
    adaptor transiently
  • reduces the ionic current
  • Magnitude of the current reduction type of
    analyte
  • Frequency of current reduction analyte
    concentration

32
Stochastic Sensors
33
a Histidine captured metal ions (Zn2, Co2,
mixture ) b CD captures anions ( promethazine,
imipramine, mixture)c biotin ligand
34
DNA sequencing
- Transmembrane potential drives translocation of
DNA or RNA through the pore - Ionic current
blockades reflect the chemical structure of
individual strands - Discrimination of different
sequences of DNA or RNA - Single resolution of
purine and pyrimidine nucleotides
Eelectrophoretically-driven translocation of a
58-nucleotide DNA strand through the
transmembrane pore of alpha-hemolysin
35
Single DNA Engineering  
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DNA? ??? ??(bead)? ??? ???? ???? ??? ??? ??????
???? ???? ?? DNA? ???? ? ??. ??? ?? ???
??????(micropipet)? ?? ????? ?? DNA ??? ???. ??
DNA? ??? ???? ?? ????DNA? ?? ??? ???? DNA?
???. ?? DNA? ????? ?? ???, CCD???? ??? ??????
???? ??.
36
Field Effect Transistor
  • A transistor is a linear semiconductor device
    that
  • controls current with the application of a
    lower-power
  • electrical signal.
  • The bipolar and field-effect transistors.
  • The bipolar transistors utilize a small current
    to control a
  • large current.
  • The field-effect transistor utilizes a small
    voltage to
  • control current The junction field-effect
    transistor.

37
In a junction field-effect transistor, or JFET,
the controlled current passes from source to
drain, or from drain to source as the case may
be. The controlling voltage is applied between
the gate and source.
With no voltage applied between gate and source,
the channel is a wide-open path for electrons to
flow. However, if a voltage is applied between
gate and source of such polarity that it
reverse-biases the PN junction, the flow between
source and drain connections becomes limited, or
regulated.
38
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39
Carbon Nanotube FET
Chen et al., PNAS, 100, 4984-4989, 2003
40
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41
Detection of biomolecular binding by using
microcantilever
  • Label-free detection of biomolecules
  • Methods for measuring the microcantilever bending
  • - Optical
  • Less amenable to monolithic
    integration and multiplexed detection
  • because of difficulties in laser
    alignment and power management
  • Interference with turbid or opaque
    fluidic and smoky environment
  • - Piezoresistive
  • Compatible with aqueous media and
    parallel cantilever arrays
  • The piezoresistors cover a large length
    of the cantilever, and high doping
  • levels are required ? the stress
    measurement is not localized
  • ? thermal and electronic noise,
    thermal drift, non-linearity in piezo-
  • response
  • More than 50 nm bending is required

42
Nanomechanical cantilever arrays
Linear position detector beam-deflection With
an accuracy of 0.1 nm
Mckendry et al., PNAS, 99, 9783-9788 (2002)
43
MOSFET-Embedded Microcantilever
  • Embedding a metal-oxide semiconductor FET(MOSFET)
    into the base of the cantilever, and recording
    decreases in drain current with deflection as
    small as 5 nm.
  • The specific biomolecular binding between ligands
    and receptors on the surface of a microcantilever
    beam
  • ? physical bending of the beam
  • ? change in the surface stress
  • ? altered channel resistance
  • ?modulation of the channel current
    underneath the gate region.
  • Embedding of MOSFET in the high-stress region of
    the cantilever to measure deflection.
  • Shekhawart et al., Science, 17,
    1592-1595 (2006)

44
N-type FET
45
Molecular imaging
  • Biomedical Sciences
  • - Ultra-sensitive imaging of biological targets
    under non-invasive in-vivo conditions
  • - Fluorescence, positron emission tomography,
    Magnetic resonance imaging
  • - Ultra-sensitive imaging
  • - Cancer detection, cell migration, gene
    expression, angiogenesis,
  • apotosis
  • - MRI powerful imaging tool as a result of
    non-invasive nature,
  • high spatial resolution and
    tomographic capability
  • Resolution is highly dependent on the
    molecular imaging agents
  • ? signal enhancement by using contrast
    agents iron oxide
  • nanoparticles

46
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47
In-vivo MR detection of cancer using NP-Herceptin
conjugates
Detection of tumors as small as 50 mg cells
48
Virus-enabled synthesis and assembly of nanowires
for lithium ion battery electrodesNam et al.,
Science, 312, 885-888, 2006
  • Smaller and more flexible Li ion batteries
  • Dimensionally small batteries Nanoparticles,
    nanotubes, nanowires as well as several assembly
    methods based on lithography, block copolymer,
    layer-by-layer deposition
  • Nanostructured materials Improvement of the
    electrochemical property of Li ion batteries
  • Monodisperse, homogeneous nanomaterials and
    hierarchical organization control are needed to
    maximize the potential

49
M13 virus
  • Helically wrapped by 2700 major coat
    proteins(p8) and minor coat proteins ( p3, p6,
    p7, and p9) around its single-stranded DNA
  • Coat proteins were used to form functional
    hetero-structured template for precisely
    positioned nano-materials

50
Predictive-based design
  • Fusion of tetraglutamate(EEEE-) to the N-terminus
    of the major coat p8 protein
  • - A general template for growing nanowires
    through the interaction of the
  • glutamate with various metal ions
  • - A blocking motif for gold nanoparticles due to
    the electrostatic repulsion,
  • reducing nonspecific gold NP binding to phage
  • Favorable interaction with the positively charged
    electrolyte polymer
  • (Ex) Design of cobalt oxide nanowires
  • - Incubation of the virus templates in
    aqueous cobalt chloride solution
  • - Cobalt oxide has large reversible storage
    capacity arising from displacement reactions
    three times larger capacity compared to
    carbon-based anodes currently used in commercial
    batteries
  • - Homogeneous and high-crystalline nanowires
    141.7 m2/g

51
Schematic diagram of the virus-enabled synthesis
and assembly of nanowires as negative electrode
materials for Li ion batteries. Rationally
designed peptide and/or materials-specific
peptides identified by biopanning were expressed
on the major coat p8 proteins of M13 viruses to
grow Co3O4 and Au-Co3O4 nanowires.

52
  • (A) TEM image of virus-templated Co3O4
    nanowires. (B) High-resolution TEM image of a
    Co3O4 viral nanowire. Electron diffraction
    pattern (Inset, upper right) confirmed that the
    crystal structure was Co3O4. (C)
    Charging-discharging curves for a virus-mediated
    Co3O4/Li half cell cycled between 3 and 0.01 V at
    a rate of C/26.5. C was defined as eight Li ions
    per hour. (D) Specific capacity versus cycle
    number for the same cell. (E) TEM images of
    differently nanostructured Co3O4 viral nanowires
    (F) TEM images of the assembly of discrete Co3O4
    nanocrystals on the p8 proteins.

Reversible capacity 600 750 mA-hour/g Twice
that of current carbon-based anodes
53
New hybrid material electrodes
  • Systematic and controlled arrangement of two
    distinct nanomaterials
  • Increase in the electrochemical properties
    through the cooperative contribution of each
    material
  • Design of composite material
  • Hybride Au-Co3O4,
  • - Gold NPs high electronic conductivity
  • - Isolation and expression of
    gold-binding peptide motif
  • (LKAHLPPSRLPS) with major coat p8
    protein
  • - Assembly of bifunctional virues expressing
    both Au- and Co3O4-
  • specific peptides with the virus coat
  • - Production and random package of two types
    p8 proteins

54
Characterization of the hybrid nanostructure of
Au nanoparticles incorporated into Co3O4
nanowires. (A) Visualization of the genetically
engineered M13 bacteriophage viruses. P8 proteins
containing a gold-binding motif (yellow) were
doped by the phagemid method in E4 clones, which
can grow Co3O4. (B) TEM images of the assembled
gold nanoparticles on the virus. (C) TEM image
of hybrid nanowires of Au nanoparticles/Co3O.
(D) Specific capacity of hybrid Au-Co3O4
nanowires. (E) Cyclic voltammograms of hybrid
Au-Co3O4 and Co3O4 nanowires at a scanning rate
of 0.3 mV/s
Specific capacity of hybride 30
greater than Co3O4 nanowires
55
  • Two-dimensional assembly of Co3O4 nanowires
    driven by liquid crystalline ordering of the
    engineered M13 bacteriophage viruses. (A and B)
    Phase-mode atomic force microscope image of
    macroscopically ordered monolayer of Co3O4-coated
    viruses. The Z range is 30 (C) Digital camera
    image of a flexible and transparent free-standing
    film of (LPEI/PAA)100.5 on which Co3O4 viral
    nanowires are assembled into nanostructured
    monolayer with dimensions of 10 cm by 4 cm. (D)
    Capacity for the assembled monolayer of Co3O4
    nanowires/Li cell at two different charging
    rates.

Linear poly(ethylene imine)/ Poly(acrylic acid)
56
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57
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