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Nano2Life 2008
Tel-Aviv University Research Institute for Nano
Science and Nano technologies
Introduction to Nano-bio-technology Interfacing
(Part 1)
Prof. Yosi Shacham-Diamand The Bernard L.
Schwartz for Nano Scale Information Technologies
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List of Content

• Introduction
• Systems concepts Sources, signals, noise,
  sensors, front-end units, signal processing,
  storage and display
• Optical signals fluorescence, bioluminescence,
  optical path design, sensors, solid state
  sensors, photon counters, system modelling
• Electrical and electrochemical sensing DC and
  impedance methods, passive electrodes based
  interfacing, nano-electrodes , active electrodes
  - Field effect transistors interfacing
• Other sensing methods magnetic sensors,
  magneto-optics, and more.
• Examples - cell on chip toxicity bio-sensors

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Introduction
Nano-bio interfacing ? how to translate the
biological information onto electrical signal ?
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Biosensors Sensing biological materials
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Biosensors Using biological components for
sensing
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Bio-receptor/ analyte complexes

• Antibody/antigen interactions,
• Nucleic acid interactions,
• Enzymatic interactions,
• Cellular interactions
• Interactions using bio-mimetic materials
• (e.g. synthetic bio-receptors).

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Signal transduction methods

1. Optical measurements - luminescence, absorption,
   surface plasmon resonance,
2. Electrochemical - potentiometric, amperometric,
   rtc.
3. Electrical transistors, nano-wires, conductive
   gels etc.
4. Mass-sensitive measurements- surface acoustic
   wave, microcantilever, microbalance, etc.).

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Biological Recognition Element
Antibody
Specificity
Enzyme
Complexity Hierarchy
DNA
Whole cell
Physiological effect
Tissue
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Bio-chips and Micro-technologies

• Micro-technologies
• Microelectronics
• Micro mechanics
• Micro fluidics
• Micro optics
• Bio-chips
• Detect biological functions
• Use biological materials or components.

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System architectures
Highlights problems
Hybrid Optimized chips Higher yield Packaging Higher cost
Fully integrated Simple process Low cost Limited design Complex process
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Systems concepts

• Sources
• signal generators,
• Noise
• sensors,
• front-end units,
• signal processing,
• storage and
• display

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Sources - signal

• Signal sources
• Optical sources fluorescence,
  photoluminescence, bioluminescence
• Electrical - Impedance spectroscopy, voltage ,
  current, electrochemical source.
• Magnetic signals magnetic beads,
• Mechanical effect Mechanical resonance,
  mechanical stress

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Sources - noise

• Noise sources
• Thermal noise due to thermal fluctuations in
  the velocity of the molecules in the electrical
  and light emitting components
• Shot noise due to variations in flux of
  particles
• Signal instability low frequency noise due to
  complex behaviors of biological material
  typical increases as the frequency decreases.
  Also calleed Excess noise or 1/f noise

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Sources - noise

• Noise sources system noise
• Fixed pattern noise spatial variations in the
  image, non uniformity,
• System electronic noise preamplifiers noise,
  sampling noise, quantization noise, analog to
  digital conversion noise,
• System mechanical noise - Microphonics
• Radio Frequency Interference (RFI)

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Front end

• Jargon the units that interface the signal
  source

Biological signal
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Fluorescence

• Biological materials using photo-biochemical
  reaction
• Photo induced luminescence photoluminescence
  example Green Fluorescent Proteins (GFP)
• Chemically induced florescence Bioluminescence
  - Example Lux
• Physical effects Luminescence from
  nano-structured materials
• Semiconductor nano-participles,

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Tunable wavelength Physical size
dependence Same chemistry
Photoluminescence from semiconductor nano
particles
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Binding DNA to Nano-crystals
Oligonucletides 18-100 bases
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Nano-crystals hybridization using DNA
Detection fluorescent microscopy
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Fluorescent Biomolecule

• Green Fluorescent Protein (GFP)from the jelly
  fish, Aequorea victoria, has served a versatile
  tool in cell biology in studying gene expression,
  protein folding and trafficking, biosensors, etc.
• With intrinsic ability to generate fluorescence
  in live tissues, using a genetically encoded tag
  by fusing GFP in-frame to a protein and the
  resulting chimeric protein was used to study
  protein function and fate in cell.

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Dependence of Excitation Spectrum on Emission
Spectrum
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Bioluminescence
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Conceptual model
Example the decomposition of four-membered ring
dioxetanones are especially relevant to
bioluminescence (Shimomura 1982).
Therese Wilson and J. Woodland Hastings,
Bioluminscence, Annu. Rev. Cell Dev. Biol. 1998.
14197230
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Example 2 A second fluorophore (F) such as GFP
is present, to which the energy carried by the
primary excited species (P) is transferred,
thereby causing this accessory fluorophore to
become excited and emit its own fluorescence.
Therese Wilson and J. Woodland Hastings,
Bioluminscence, Annu. Rev. Cell Dev. Biol. 1998.
14197230
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Other optical methods

• Surface Plasmon Resonance (SPR)

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Surface Plasmon Resonance (SPR)
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System architectures
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System architectures

• Chips flat platforms, sensors below or above
  the chip

fluorescence detection of Cy5-labeled
Streptavidin using a 4X4 photodiode array IC
biochip. Excitation by a 12 mW HeNe laser (632.8
nm).
T. Vo-Dinh et al. , Sensors and Actuators, B 74
(2001) 2-11
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Schematic diagram of an integrated DNA biochip
system
Vo-Dinh T, Alarie JP, Isola N, Landis D,
Wintenberg AL, Ericson, MN (1999) Anal Chem 71
358363
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Bio-chips

• Portable,
• low cost in high volumes,
• low power,
• can be integrated with other components

Chii-Wann Lin et al, DEVELOPMENT OF MICROMACHINED
ELECTROCHEMICAL SENSOR AND PORTABLE METER SYSTEM,
a Proceedings of the 20th Annual International
Conference of the IEEE Engineering in Medicine
and Biology Society, Vol. 20, No 4,1998
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System architectures

• Fiber optics based

Detection of a Benzoapyrene tetrol (BPT), a
biomarker.
T. Vo-Dinh et al. , Sensors and Actuators, B 74
(2001) 2-11
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Sensors

• Optical sensors
• Solid state devices
• photodiodes, Avalanche photodiodes,
• Photoconductors
• Sensor arrays CMOS imagers, Charge Coupled
  Devices (CCD)
• Photomultipliers
• Electrical sensors
• Passive components
• Electrodes,
• Conductive materials
• Active components
• Transistors
• Semiconductors based
• CNTs

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Single detectors vs. Vectors and arrays
Single
Vector
Array
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Optical sensors
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Integrated Solid state detectors

• Single detectors photodiodes, avalanche
  photodiodes, photomultipliers
• Detector arrays CMOS imagers, CCDs

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Discrete detectors

1. Single container
2. Large volumes
3. Can be optimized electronics area is not
   limited by the biochip

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Photomultipliers converts photons flux to
electrical current with high internal gain

• Highlights
• Very sensitive
• Problems
• Slow
• Expensive
• Difficult to integrate

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Photodiodes converts photon flux to current

• Highlights
• Si Integrated Circuit compatible
• Low cost
• Fast
• Problems
• Less sensitive than photomultiplier

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Advanced diode based sensors

• APD - Avalanche Photo Diodes internally
  amplified by the avalanche multiplication process
• Another versions is the SPAD - Single Photon
  Avalanche detector

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Detector arrays

• Highlights
• Many containers battery on a chip
• Can be software controlled for the collection
  area
• Problems
• Electronics area is limited by the biochip
• Complex optical constraints

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Example integrated photodiode with CMOS circuit
Micro-luminescent detection ASIC Chip area 2mm
X2mm Photodiode area 1.47 mm2
Michael L. Simpson et al., Sensors and actuators
B, 2002, 179-185
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Electrical sensing

• Electrochemical
• Redox reactions
• Electrical
• SAM-FET
• Nano wires
• Conducting Electro-active Polymers

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Single cell electrical sensors
Detection method Detection range/resolution Metabolism
Calorimetric, ISFET, Electrochemical, PEBBLEs, SECM 0-280 mM / 0.1 mM Oxygen
Calorimetric, ISFET, Electrochemical 0.1-10 mM / 0.1 mM CO2
Calorimetric, Luminescence, ISFET, Electrochemical, LAPs 5.2-8.5 mM / 0.1 pH
Calorimetric, Luminescence, ISFET, Electrochemical, LAPs, SECM 3-23 mM Glucose
Calorimetric, Luminescence 0-5 mM ATP
Luminescence 0-125 mM NADH, FADH2
Yotter and wilson, monitoring metabolic activity
in single cellspart ii nonoptical methods and
applications, IEEE sensors journal, vol. 4, no.
4, august 2004
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Electrical sensors

• FET based methods FET Field Effect
  Transistors
• ISFET Ion Sensitive FET
• CHEM-ET Chemically Sensitive FET
• SAM-FET Self Assembly Monolayer Based FET
• PEBBLEs Probe Encapsulated by Biologically
  Localize Embedding
• LAPS Light Addressable Potentiometric Sensors
• SECM - Scanning Electro Chemical Microscopy

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Electrochemical sensors

• Electrode arrays sensing potential, charge
• Electrochemical probing using electro chemical
  reaction for sensing

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Electrode options
Highlights Problems
Metallic nano-electrodes - Simple - Standard circuits - Single cell or 1D array only - Stability - Small signal - 1 circuit/channel
SAM-FET / CMOS compatible - Large signal - Single cell - 2D array possible - Stable - Small area - Complicated - Dedicated circuits
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Microelectrodes
Ring-type interdigitated Electrodes
Nanoelectrodes
ZHU AND AHN ELECTROCHEMICAL DETERMINATION OF
REVERSIBLE REDOX SPECIES AT IDA
MICRO/NANOELECTRODES, IEEE TRANSACTIONS ON
NANOBIOSCIENCE, VOL. 4, NO. 2, JUNE
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Carbon nanotube electrodes
Yael Haneins group, school of electrical
engineering, TAU
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Massive amount of interconnected ordered cell
clusters Yael Haneins group, school of
electrical engineering, TAU
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MOSFET based sensing
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Micro- pH and Bio Sensors Based on Field Effect
Transistors
On-chip devices aiming for high accuracy
bio-molecules detection
Achievements
?Formation of monolayer patterns
?Biomolecule immobilization onto the monolayer
patterns
BSA
DNA
Field Effect Transistors, etc.
J. Phys. Chem. B 108, 3240 (2004) Chem. Lett.
33, 176 (2004) Chem. Lett. 33, 284 (2004)
Jpn. J. Appl. Phys. 43, L105 (2004) Sens.
Actuators B, in press.
Future Prospects
?Sensing of bio-related materials (SNPs,
Analyses of proteins and cells)
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Urease Detection with New FET Sensor
Urease
NH2CONH2 3H2O
HCO3- 2NH4 OH-
pH shift
(D. Niwa et al. submitted to Sens. Actuator B)
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Nano-bio interfacing
Nano-wires interfacing
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Nano-wire Field Effect Transistors (FET)

• P-type devices
• Hole mobility 1500 cm2/V.sec.

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Nanowires Biosensors

• Functionalize the nano-wires
• Binding to bio-molecules will affect the
  nano-wires conductivity.

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Example Specific protein sensing in
Functionalized Nano-wires
Cui et al, Science 29, 1298, 2001
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Conducting Electro-active Polymers
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Chemical and Biological Sensors Based on
ElectrochemicalDetection Using Conducting
Electro-active Polymers
Immobilization of biotin to IME/PPy-hydrogel
device. Reaction of surface-available, N-acid
moieties of the pyrrole/4-(1-pyrrolyl) butaric
acid polymer conjugate with 5-(biotinamido)pentyl
amine using carbodiimide linking chemistry
produces a biotinylated copolymer at the surface
M. Brahim et al, Microchim. Acta 143, 123137
(2003)
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Chemical and Biological Sensors Based on
ElectrochemicalDetection Using Conducting
Electro-active Polymers
Principle of operation of the polypyrrole-based
conductimetric biosensor for glucose based on
strong biotin-streptavidin affinity. Enzyme
glucose oxidase
M. Brahim et al, Microchim. Acta 143, 123137
(2003)
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Chemical and Biological Sensors Based on
ElectrochemicalDetection Using Conducting
Electro-active Polymers
(A) Response profiles of the glucose-responsive
PPy-based conductimetric biosensor toward
different glucose levels as measureusing the
EPSIS analytical methodology
(B) the change in initial conductivity of the PPy
membrane (slopes) after the first few seconds of
the enzymatic reaction
M. Brahim et al, Microchim. Acta 143, 123137
(2003)
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Magnetic sensing

• Actually it is using magnetic fields to sense
  magnetic nano-particles that have been attached
  to biological molecules.

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Surface coverage of the modified magnetic
particle and substrate
Table
Surface coverage
APTES
Streptavidin
Magnetic particle
18 1)
66 3)
100 (Tightly packed) 2)
Substrate
12 4)
Surface coverage (The area modified with
molecules) / (The total surface area of the
substrate or particle) x 100.

1. APTES numbers were determined by photometric
   method using sulfo-LC-SPDP.
2. This was evaluated by ellipsometry and XPS
   analysis.
3. Streptavidin numbers on a particle were obtained
   by determining protein amount.
4. Streptavidin numbers were calculated by
   determining fluorescence labels conjugated with
   the streptavidin.

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Stabilization of Magnetic Particles with various
Streptavidin Conc.
Magnetic nano -particle
Streptavidin
10 min reaction
100 mg/ml
1 ng/ml
100 ng/ml
None
Cleaning
5 min deposition
Biotin-stabilized substrate
T. Osaka, 2006
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Stabilization of Magnetic Particle vs.
Streptavidin Concrntration
(A)no addition
(B)1 ng/ml
(C)100 ng/ml
(D)100 mg/ml
Particles are stabilized with the reaction of
biotin-streptavidin
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MFM Observation of Stabilized Magnetic Particles
on Substrate
A
1 mm
SEM image
B
C
AFMimage
MFM image
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Magnetic sensing
Sensing plate
Magnetic head
Activated pixel
Idle pixel
Magnetic field sensor
Magnetic sensing
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Magnetic-optic sensing
Sensing plate
Polarization Detector
Optics
Light source
Activated pixel
Idle pixel
Polarizer
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Schematic diagram showing the components of
the sandwich immunoassay for CRP.
Phase locked loop
CRP - C-reactive protein
PMP Para Magnetic Particle
Reader layout
Richard Luxton et al, Anal. Chem. 2004, 76,
1715-1719
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Schematic illustration of biotin streptavidin
reaction on SAM-modified substrate. Biotin is
attached to the APS-patterned substrate.
Streptavidin-modified particles are injected into
a channel on a glass plate. The particles are
bound by a specific interaction between biotin
and streptavidin.
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Mechanical sensing Cantilever based sensing
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Bio-molecule sensing
Detection of biomolecules by simple mechanical
transduction - cantilever surface is
covered by receptor layer (functionalization) -
biomolecular interaction between receptor
and target molecules (molecular
recognition) - interaction between adsorbed
molecules induces surface stress change ?
bending of cantilever
target molecule
receptor molecule
gold
SiNx cantilever
target binding
deflection ?d
B. Kim et al, Institut für Angewandte Physik -
Universität Tübingen
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A cantilever as a mass-sensitive detector
f1
f1
?m
f2
f2
A mass sensitive resonator transforms an
additional mass loading into a resonance
frequency shift ? mass sensor
B. Kim et al, Institut für Angewandte Physik -
Universität Tübingen
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Surface Stress induced bending
? ? surface stress change t thickness of the
beam L length of the beam E Youngs modulus
of the material ? Poisson ratio of the
material ? d deflection of the end of the beam
Stoney formula
Cantilever bending can potentially detect single
molecules, however they are noise limited
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Optical detection of analyte binding
Detection scheme
B. Kim et al, Institut für Angewandte Physik -
Universität Tübingen
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Thank You! ?