Nanowire Sensing Systems - Nano-Tera

Transcription

05.02.2016
Nanowire Sensing Systems for
Biomedical Applications
Prof. Dr. Sven Ingebrandt
University of Applied Sciences Kaiserslautern –
Campus Zweibrücken, Germany
Prosense Winter School @ EPFL Lausanne, 04.02.2016
www.hs-kl.de
Ingebrandt | Kaiserslautern | 02. February 2016 | 1
Outline
 Introduction to the University of Applied Sciences Kaiserslautern
 Motivation
 Silicon nanowire (SiNW) detection principles
 Top-down fabrication of SiNW sensors
 Electronic readout principles for SiNW sensors
 Electrochemical characterization
 Bioassays with SiNW sensors :
1.
2.
3.
4.
5.
DNA immobilization and hybridization detection
Ca2+ ions – peptide interaction
Immunoassays
Detection of cellular signals
Impedimetric detection of cellular adhesion
 Possible future assays with SiNW sensors
 Summary
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05.02.2016
Introduction to the University of
Applied Sciences Kaiserslautern
Introduction to the UASK: Zweibrücken
Area: 70.64 km2
Population: 34,109
City of roses and horses
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05.02.2016
Introduction to the UASK
 Spread over three cities Kaiserslautern, Pirmasens, and Zweibrücken
 5 faculties, 23 courses, 6000 students, ~168 professors
Campus Kaiserslautern
 Applied engineering sciences
 Construction and design
Campus Pirmasens
 Applied logistics- and polymersciences
Campus Zweibrücken
 Economics
 Informatics and
Microsystemtechnology
Introduction to the UASK: Study in Zweibrücken
Speciality: Large clean room (300 sqm) for education and research
Student courses / Summer schools
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05.02.2016
Introduction to the UASK: Nanoimprint Lithography
Installed in cleanroom of FHKL
DFG-‘Large equipment proposal:
Obducat Eitre 6“
Nanoimprint-Lithography
Introduction to the UASK: Schottky Field Emission SEM
Zeiss Gemini Supra with high
Resolution EDX
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Introduction to the UASK: Raman/AFM spectrometer for TERS
Horiba LabRAM system combined with AFM for high
resolution Raman spectroscopy
Biomedical Signalling Research Group (AG Ingebrandt) – December 2016
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Motivation
Bioelectronics = Biology + Electronics
+
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Activities of the research group Biomedical Signalling
Motivation: Biosensors and applications
Applications
Ideal properties:
• Food analysis
• High sensitivity
• Drug development
• High selectivity
• Criminal investigation
• Cost effective devices
• Medical diagnosis
• Real-time detection
• Fast response time
• Test of donated organs
http://www.minimed.com
• Point-of-care usage, …
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05.02.2016
common source
Field-Effect Transistor
common source
gate
200 µm
20 µm
drain
Fabrication of ISFET chips to interface individual cells
Former
FET
generation
IMM, Mainz
MPIP Mainz
FZJ Jülich
•
•
New
FET
generation
UASK
Zweibrücken
ISFET sensors were fabricated
having almost flat topography
except for transistor gate
openings.
Cytotoxic T cells: Important role in
the immune system by migrating
throughout the whole body,
recognizing, and then eliminating
specific pathogens.
J. K. Y. Law, et al., Human T cells monitored by impedance spectrometry
using field-effect transistor arrays: A novel tool for single-cell adhesion
and migration studies, Biosensors and Bioelectronics, 67, (2015) 170-176.
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Electronic Cell-substrate Adhesion studies with Human T-killer cells
2.00
Normalised Transfer Function (H)
1.75
5 min
10 min
15 min
20 min
25 min
30 min
1.50
1.25
1.00
0.75
0.50
10k
100k
1M
Frequency (Hz)
Adhesion
Adhesion
•
•
Transfer function was changed over time (30
min)
•
The attachment and detachment can be
observed clearly
A significant difference in transfer function
between cell-covered and cell-free gate
depending on type of doping molecules
J. K. Y. Law, et al., Human T cells monitored by impedance spectrometry
using field-effect transistor arrays: A novel tool for single-cell adhesion
and migration studies, Biosensors and Bioelectronics, 67, (2015) 170-176.
Individual migrating-activated T cells monitored by field-effect transistors
Transfer function (H)
3.00
2.75
2.50
2.25
Chip 1 (Ch 13)
2.00
0
500
1000 1500 2000 2500 3000 3500 4000
Time (s)
Adhesion and activation studies of primary
human T-killer cells (cooperation with M. Hoth –
University Saarland – Homburg)
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Detection of signalling molecules involved in cancer signalling
Specific binding reaction on nanowires
Stern, E.; Vacic, A.; Li, C.; Ishikawa, F.N.; Zhou, C.W.; Reed, M.A.; Fahmy, T.M. A nanoelectronic enzyme-linked
immunosorbent assay for detection of proteins in physiological solutions. Small 2010, 6, 232–238.
Comparison of ISFET dimensions
Burj Kalifa
828 m
Does Size Matter?
Statue of Liberty
46 m + 17 m socket
Giraffe
6m
2400x5 µm2
200x10 µm2
Y. Ishige, et al., Biosens
H.J. Park et al., FEBS
Bioelectron 24,1096, (2009). Letters 583, 157 (2009).
8x4 µm2
ISFET design
(© AG Ingebrandt,
2015) for individual
cell interfacing
Beluga
sturgeon
1.4 m
Three french
baguettes
0.8 m – small
diameter
0.08x4 µm2
0.005x1 µm2
Y. Cui et al., Nano
SiNW-FET design
(© AG Ingebrandt, 2015) Letters 3, 149 (2003).
for biosensing
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Motivation: ISFET-based biosensors – General principle
• Label – free and real-time detection
• Direct electronic readout signal
• Possibility of a miniaturization
𝑉𝑇𝐻 = 𝐸𝑟𝑒𝑓 − 𝛹𝑠 + 𝜒𝑠𝑜𝑙 −
Transfer characteristics
𝛹𝑆𝑖 𝑄𝑜𝑥 − 𝑄𝑠𝑠 𝑄𝐵
−
−
+ 2𝛷𝐹
𝑞
𝐶𝑜𝑥
𝐶𝑜𝑥
Motivation: Silicon nanowire biosensor
Advantages
• Ultrahigh surface-to-volume ratio
• Dimensions of the biomolecules are
comparable to that of the nanowires
• Strong influence of the surface effects
to the electronic properties
High sensitivity
• Possibility to create dense arrays
Requirements
• Wires in the 10 nm range with good carrier
mobilities
• Low density of trap states at the interfaces
(high subthreshold swing)
• High transconductance
• Stability and reliability of the output signal
• Processability of the sensors
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05.02.2016
Biomolecules binding to silicon nanowires
The Debye screening problem – Fundamental Limitation of the
typically-used potentiometric approach
**
*
When potentiometric sensing is done, almost no sensitivity towards
biomolecules charges is left.
DC method works good for smaller molecules and for short DNA strands.
* W. Huang, A. K. Diallo, J. L. Dailey, K. Besara, and H. E. Katz, Electrochemical processes and mechanistic aspects of field-effect sensors for biomolecules, J.
Mater. Chem. C, 2015, 3, 6445-6470.
** E. Stern, R. Wagner, F. J. Sigworth, R. Breaker, T. M. Fahmy, and M. A. Reed, Importance of the Debye Screening Length on Nanowire Field Effect Transistor
Sensors, Nano Lett., 2007, 7, 3405–3409.
AC – Transfer Function Impedance Spectroscopy – Impedimetric detection
Detection principle: Sensor is brought into working point by setting VDS and VGS just like in timedependent DC mode. Additionally a small sinusoidal test voltage Vstim is applied to the reference
electrode. This is scanned over frequency. The system nanowire plus its first amplifier stage is acting
like a frequency low pass. The spectrum can be described by a transfer function (mathematical
representation of the electrical circuit). In the spectrum the impedance of the biomolecular layer is
hidden.
This technique can be applied by lock-in amplifier which is boosting the signal-to-noise ration
tremendously.
Transfer Function:
Vstim ( j )
Vstim
H ( j ) 
Vout ( j )
Vstim ( j )
Vout ( j )
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Detection principles: Impedimetric measurements
ISFET as impedimetric biosensor – Transistor-Transfer Function (TTF)
Vstim()
Transfer Function:
H ( ) 
Vout ( )
Vstim ( )
Vout()
Low cutoff frequency
 1  Rmem (Cmem  Cox )  RmemCox
 2  RmemCmem
High cutoff frequency
Schasfoort RBM et al., (1989) Sensor Actuator, 18, 119
Kharitonov, AB et al., (2000), Sens. Actuator B-Chem., 70, 222
Antonisse, MMG et al., (2000), Anal. Chem., 72, 343
Experimental setup
• Characterization of the FET
devices
• Measurement of the
frequency-dependent
transfer functions
• Recording of the timedependent data
Advantages:
• Measurement of the impedance spectra up to 50 MHz
• Time-dependent readout at several different frequencies, simultaneously.
A. Susloparova, et al., Electrical Cell-substrate Impedance Sensing with field-effect transistors is able to unravel cellular adhesion and detachment
processes on a single cell level, Lab on a Chip, (2015), 15, 668-679.
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05.02.2016
Bio-sensing applications: Electronic DNA-chip with micro-ISFETs
• Novel, lock-in based signal
transduction concept
• Reliable recording
• Fast sampling
Frequency (Hz)
S. Ingebrandt, Y. Han, F. Nakamura, A. Poghossian , M.J. Schöning, and A. Offenhäusser.
Label-free detection of single nucleotide polymorphisms utilizing the differential transfer function of field-effect transistors,
Biosensors and Bioelectronics, 2007, 22, 2834–2840.
Top-down fabrication of
SiNW arrays
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05.02.2016
Dr. X.-T. Vu, R. Stockmann
Fabrication process of Si-NW arrays
 First trial: Electron beam lithography
Project start in 2006
At project start: Si nanowire by e-beam lithography and RIE (in cleanroom of IBN-2)
- Electron Beam Writer Leica EBPG 5000 Plus
- High resolution (down to 5nm)
- Maskless patterning
Bad surface quality
Strong hysteresis
Slow process
 Silicon nanowire device concept:
 Reed group (Yale)
 Top-down approach
 Biomolecular sensing demonstrated
[Source: Stern er al, Nature, February 2007]
Wet etching process with tetra methyl ammonium hydroxide (TMAH)
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05.02.2016
Wafer-scale fabrication of SiNW arrays
SiO2
Si
X.T.Vu et al, Physica Status Solidi, 206 (2009) 426
X.T. Vu et al, Sensors and Actuators – B Chemical, 144 (2010) 354
Buried oxide
(BOX)
Chip designs
4×4 Arrays
6 wires parallel
Length: 3mm
Width: 100, 200, 500, 1000nm (mask
design)
pitch: 200mm
X.T.Vu et al, Physica Status Solidi, 206 (2009) 426
X.T. Vu et al, Sensors and Actuators – B Chemical, 144 (2010) 354
28×2 Arrays
Single wires
Length: 10mm, 20mm, 40mm
Width: 200, 400nm (mask design)
pitch: 50mm or 10mm
Integration with micro-fluidics
16×16, 32×32 and 128×128 arrays
X.T. Vu, et al.,Phys. Status Solidi A-Appl. Mat., 207, 4, (2010) 850-857.
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05.02.2016
Fabrication of larger Si-NW FET arrays
Two designs:
16×16 with 100 µm pitch (256 channels)
32×32 with 50 µm pitch (1024 channels)
Fabrication process of Si-NW arrays: SiNW devices
• Wafer-scale, reproducible process
Base: 120 nm
• Size of SiNW can be as small as 60 nm at the bottom (in current process)
• High quality of the gate oxide and smooth surfaces
• High quality of the passivation layer – stable operation in electrolyte solution
Top: 67 nm
Length: 40000 nm
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05.02.2016
Fabrication process of Si-NW arrays:
Wafer-scale processing
28x2 arrays
4 inch wafer
128x128 arrays
32-channel SiNW sensor chip design
 32 single NW FET devices with dip-chip configuration.
NW dimensions:
 250nm(width) by 12µm (length), 60nm height
Si(doped contact lines) Metallic contact lines
8 sets of 4
NWs
separated by
250µm
7mm
1 set of 4 NWs
Source
5µm
10mm
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05.02.2016
Residual resist
Common Source
Individual Drains
Process flow for SiNW fabrication
1µm
Fabricated SiNW devices
A
200 µm
D
C
5 µm
100 µm
B
6.79 K X
5.00 kV
200 nm
174.5 K X
5.00 kV
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05.02.2016
Portable amplifier for the FET arrays
• Simultaneous measurement of all 16
channels
• Precise temperature control
• Amplifier stages with 1x, 10x, 30x, 100x
• Differential readout with reference channel
• Measurement of the transfer function by a
16x lock-in circuit
• Portable system with USB-interface to PC
• Custom-made software (Delphi 5.0)
250 kW
• Sampling rate 1 Hz (up to 10 kHz per
channel with external USB-DAC card)
Dr. X.-T. Vu
Fabrication process of Si-NW arrays: Chip packaging
•
Wire – bonding
• Contact isolated: Medical epoxy, PDMS
• Can be used directly or integrated with
micro-fluidics
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05.02.2016
T. C. Nguyen, Dr. X.-T. Vu
NEW 32-channel readout system design
•
•
•
Here is the design for one channel, different channels are currently addressed by
switching
The system is flexible for different ranges of drain-source currents
Suitable for both characterization and impedimetric measurement of FET devices
T. C. Nguyen et al., Physica Status Solidi (a), 210, 5, 870–876 (2013).
NEW amplifier system
LabVIEW programming
 State machine programming:
 Program is divided into
different states
 Easier to read
 Easier to manage
 Easier to scale
 Stand-alone application: no
LabVIEW development tool is
needed
Headstage setup
T. C. Nguyen et al., Physica Status Solidi (a), 210, 5, 870–876 (2013).
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05.02.2016
NEW amplifier system
32 channel system for SiNW
recordings
DC and AC capability
Electrochemical characterization
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05.02.2016
Electrical characterization setup
•
Understanding the SiNW electrical transport properties
•
Finding of a stable configuration for biosensor experiments
Back - gate characteristics: IDS(VBG)
Front gate in air:
Front gate with electrolyte (floating potential or applied voltage)
Front - gate characteristics: IDS(VFG)
Back gate floating
Back gate with applied voltage
Dr. X.-T. Vu
Electrical characteristics
Implanted CL
Non-implanted CL
Back-gate control
p-type only
p- and n-type
(in dry and wet
unstable operation
unstable operation
Front-gate control
p-type only
n-type only
(in wet environment)
stable operation
stable operation
environment)
• Front-gate configuration was identified for optimum biosensing experiments
• In any case, an electrochemical reference electrode is needed for the front gate contact
to keep the system stable and the readout signal reliable
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05.02.2016
Dr. X.-T. Vu
Electrical characteristics
Front-gate transfer characteristics: IDS(VGS)
𝑆=
𝑘𝑏 𝑇
𝐶𝐷
ln(1 +
)
𝑞
𝐶𝑜𝑥
• P- enhancement and long-channel transistor
• VTH = - 0.8 V at pH 7
• gm, max (at VDS=-2V): 0.5 µS to 10 µS
• Ion/Ioff =105-106
• S = 80 - 110 mV/decade, size dependent
Dr. X.-T. Vu
Electrochemical characterization
pH sensitivity: Two possible operation modes
Saturation region
Sub-threshold region
X.T. Vu, et al.,Phys. Status Solidi A-Appl. Mat., 206, 3, (2009) 426-434.
pH sensitivity : 42 mV / pH and does not depend on size and operation mode
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05.02.2016
Bioassays
Assay 1: DNA detection
Dr. X.-T. Vu
Assay 1: DNA immobilization and hybridization
I
II
150 mV
III
SiNW
Immobilization: ΔVG up to 150 mV
Hybridization: ΔVG up to 40 mV
40 mV
ISFET
Immobilization:
Hybridization:
ΔVG = 1-10 mV
ΔVG = 1-4 mV
Han et al. 2006
SiNW sensor is much more sensitive than our microscale ISFETs
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05.02.2016
Dr. R. GhoshMoulick, Dr. X.-T. Vu
Assay 1: Covalent attachment of DNA probes
Protocol (a)*:
• Silanization with 3-aminopropyltriethoxysilane
• Usage of succinic anhydride as cross linker
• Covalent attachment of amino-functionalized
DNA
Protocol (b)**:
• Silanization with 3-glycidoxypropyltrimethoxysilane
• Direct, covalent attachment of amino-functionalized
DNA and of proteins
* Han et al., 2006; Ingebrandt and Offenhäusser, 2006.
**Ghosh-Moulick et al., 2009.
M. Schwartz, Dr. X.-T. Vu
Assay 1: Surface modification - silanization
Gas phase silanization
 Homogenous monolayers
 Encapsulated material not
damaged
Contact angle measurement
 Quantify wettability
 Qualify silanization:
 Did silanization work?
 Is layer homogeneous?
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05.02.2016
Assay 1: DNA microspotting & electrical readout
 Site-specific spotting of different capture
molecules
 Differential read-out
possible
 Defined distances to
200 µm
avoid cross-contaminations
reference
electrode
electrode
holder
Source
Nanowire
Drain
Assay 1: Surface modification steps - overview
3-Glycidoxypropyltrimethoxysilane (GPTES)
 No cross-linker due to covalent binding
 Creation of monolayers
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05.02.2016
Assay 1: DNA detection – dc readout
Results unpublished
Applied concentrations:
 1 µM capture DNA
 0.5 µM cDNA
 1 µM cDNA
~ 200 mV
~ 35 mV
~ 50 mV
~ 75 mV
VTH shifted to the right due to binding of negatively charged DNA molecules
Assay 1: DNA detection – dc readout
Why are smaller wires more sensitive?
Two effects of ‘conductivity change’
Miriam Schwartz, et al., DNA detection with top-down fabricated silicon
nanowire transistor arrays in linear operation regime, physica status solidi
(a), (2016) accepted
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05.02.2016
Assay 1: DNA detection – ac readout
Exemplary measurement
0.5 µM cDNA
GPTES
capture
DNA
block
1 µM cDNA
Bioassays
Assay 2: Ca2+ ions – peptide interaction
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05.02.2016
M. Hitzbleck, Dr. X.-T. Vu
Assay 2:
Ca2+
ions – peptide interaction
M. Hitzbleck et al. Submitted to Langmuir
M. Hitzbleck et al., Physica Status Solidi (a), 210, 5, 1030–1037 (2013).
Bioassays
Assay 3: Immunoassays
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05.02.2016
Assay 3: Antibody-antigen detection by SiNW (Human CNTF*)
CNTF is known to alter the self – renewal process of
neuronal stem cells and the progenitor cell division and
differentiation
It is used in the therapy and gene-therapy of retinal ganglion
cell damage or loss
CNTF seems to have a protective role in multiple sclerosis
mouse model regarding demyelinisation and plays a role in the
pathogenesis of other neurodegenerative diseases
CNTF is a good model system for the future measurements
of other neurotropic factors such as BDNF, GDNF, FGF and
NGF
CNFT is usually detected by an elaborated ELISA method
Sensitivity of the ELISA method is 30pg/mL at a dynamic
range of 30 pg/mL - 3000 pg/mL
*CNTF: Ciliary neurotrophic factor
Brain-Derived Neutrophic Factor (BDNF)

Polypeptide (27 kDA)

IP = 9.01

Stimulates neuronal development, growth,
survival, function in CNS & PNS

Influences neuronal network
communication
A test system with a lower limit of
detection than ELISA is needed!
B. K. Pedersen, Exp. Physiol. 94 (2009) 1153-1160.
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05.02.2016
Assay 3: Antibody-antigen – dc readout
Results unpublished
 1 µg/ml capture antibody
 Block with 1 % BSA

 5 pg/ml BDNF
 10 pg/ml BDNF
 50 pg/ml BDNF
1 pg/ml BDNF
0.001 x PBS
~ 50 mV
~ 90 mV
~ 110 mV
~ 120 mV
~ 130 mV
VTH shifted to the left
SiNW FET - dc-readout of BDNF
1.)
2.)
3.)
4.) – 7.)
Exemplary DC measurement
Stepwise recordings
1.) GPTES
2.) capture ab
3.) block
4.) 1 pg/mL BDNF
5.) 6 pg/mL BDNF 6.)
16 pg/mL BDNF
7.) 66 pg/mL BDNF
n = 11
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SiNW FET - AC-detection of BDNF
1.)
2.)
4.) – 7.)
3.)
Exemplary AC measurement
Stepwise recordings
1.) GPTES
2.) capture ab
3.) block
4.) 1 pg/mL BDNF
5.) 6 pg/mL BDNF 6.)
16 pg/mL BDNF
7.) 66 pg/mL BDNF
n = 11
Comparison of the silicon nanowire data with the golden standard
ELISA test
Collected results from all tests…
SiNW DC recordings
SiNW AC recordings
10
0.88
0.78
0.58
0.48
0.1
0.38
AC signal (µS)
1
SiNW data
0.28
0.18
0.01
1
10
100
BDNF concentration (pg/ml)
1000
1
2
1
10
100
1000
0.1
Optical density
20
0.68
Optical density
DC signal (mV)
10
ELISA data
0.2
ELISA kit
ELISA kit
0.02
0.01
BDNF concentration (pg/ml)
In our trials the silicon nanowire platform is about 20-fold lower in limit
of detection than the golden standard!
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05.02.2016
Dipti Rani
Problem of non uniform surface modification while micro spotting
1 µM
500 nM
Typical microspotting result – nonideal surface coating of DNA
Structures to examine the local
variation of SiNW signals
Electrical Characterization of SiNWs
-8
2.0x10
Drain source current(A)
0.0
-8
-2.0x10
-8
-4.0x10
-8
-6.0x10
-8
-8.0x10
-7
-1.0x10
-7
-1.2x10
-7
-1.4x10
-1.0
-0.8
-0.6
-0.4
-0.2
0.0
Gate source voltage(V)
0.40
Negative threshold Voltage(Vth)
0.35
32-channel SiNW chip on carrier
0.0
Drain current / A
Drain current / A
0.0
-500.0n
-1.0µ
Vg=0V
Vg=-0.5V
Vg=-1V
Vg=-1.5V
Vg=-2V
-1.5µ
-2.0
-1.5
-1.0
-0.5
Drain Source bias / V
-2.0µ
- 0.5
-3.0µ
- 1.0
-4.0µ
-5.0µ
- 1.5
0.25
0.20
0.15
0.10
0.05
-6.0µ
-7.0µ
0.0
Vds = 0
-1.0µ
0.30
- 2.0
0.00
-3.0
-2.5
-2.0
-1.5
-1.0
-0.5
0.0
Gate voltage at Ag-AgCl reference / V
SiNW act as long-channel ISFET devices
0
5
10
15
20
25
30
Channel Number
Vth variation of different channels on a single chip
Average Vth=-0.315 ± 0.023 V
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05.02.2016
Biosensing with SiNW-FET devices
0.0
pH5
pH6
pH7
pH8
pH9
-20.0n
-40.0n
-60.0n
-80.0n
-100.0n
-120.0n
-140.0n
-1.0
-0.8
-0.6
-0.4
-0.2
0.0
Threshold Voltage change /  Vth
Drain source current / A
pH sensing with SiNW-FET array
Average Vth = 43 +/- 3 mV.pH
-1
0.15
0.10
0.05
0.00
4.5 5.0 5.5 6.0 6.5 7.0 7.5 8.0 8.5 9.0 9.5
pH value of buffer solution
Effect of silanization:
600.0m
Vds = 100 mV
Threshold voltage / V
Drain current / A
0.0
-20.0n
-40.0n
-60.0n
-80.0n
-100.0n
-1.0
-0.8
-0.6
-0.4
-0.2
0.0
Silicon nanowire
Silicon nanowire functionalized with GPTES
500.0m
400.0m
300.0m
200.0m
100.0m
0.0
0
5
10
15
20
25
30
-th Si nanowire ISFETs on chip
PSA aptamer & binding mechanism

Some NW sets functionalized with receptors (PSA aptamer) and others with
control using microspotting technique.

PSA binding with aptamers
0.0
-8
-2.0x10
-8
Drain source current(A)
-4.0x10
-8
-6.0x10
-8
-8.0x10
-7
-1.0x10
-7
GPTES
Aptamer
Ethanolamine
PSA 30nM
-1.2x10
-7
-1.4x10
-7
-1.6x10
-7
-1.8x10
-7
-2.0x10
-7
-2.2x10
-1.0
-0.9
-0.8
-0.7
-0.6
-0.5
-0.4
-0.3
-0.2
-0.1
Gate source voltage(V)
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05.02.2016
Fluidic encapsulation of SiNW chips
Contact pads
32-channel SiNW-FET chip
(size 7x10 mm2)
32 SiNW sensors
Fluidic cell
(made from PDMS)
Bioassays
Assay 4: Detection of cellular signals
36
05.02.2016
Dr. X.-T. Vu
Assay 4: Recording of extracellular action potentials
Primary cardiac myocytes (E18) and HL-1 cells were cultured on NW-FETs arrays
Once a confluent layer is formed cells are
spontaneously beating.
Rat cardiac myocytes at 5 DIV
Signals can be site-selectively recorded by
the NW-FET array.
Dr. J. F. Eschermann, Dr. X.-T. Vu
16-channel recording of action potentials
HL1 - cell line
Signals show a similar shape as compared to ISFETs
Signal-to-noise ratio is improved compared to ISFETs
J. F. Eschermann et al., Applied Physics Letters, 95, 083703 (2009)
37
05.02.2016
Dr. X.-T. Vu
Results unpublished
Primary cardiac myocyte culture!!!
Each of the 16-channels is responding
High signal-to-noise ratio : 5 – 10 due to the strong coupling between the
cells and the SiNWs
Bioassays
Assay 5: Impedimetric detection of
cellular adhesion
38
05.02.2016
Dr. X.-T. Vu
Assay 5: Impedimetric detection of cellular adhesion
Vstim
1Hz – 1MHz
IDS
Dr. X.-T. Vu
Results unpublished
HEK cell-SiNW coupling: Detailed SEM analysis
39
05.02.2016
Dr. X.-T. Vu
Results unpublished
Future directions
40
05.02.2016
D. Rani, Dr. X.-T. Vu
New research projects related to SiNW sensing
Developing a NW-based sensor platform for detection of PCa in parallel with
fluorescence techniques
Fluorescent dye
Analyte
Receptor
Linker
D. Rani, Dr. X.-T. Vu
New research projects related to SiNW sensing
Si NW FET Fabrication on Sapphire substrate for combined optical
and electrical sensing
 Dip Chip:
 32 single NW devices on 7*7mm 2 chip.
 NW dimensions- 1µm(length)*0.1µm(width).
 Common source configuration.
Si NWs
Drain
Sapphire substrate
41
05.02.2016
Case study: AC – For sensing beyond Debye Limit
Detection principle: The detection of charged biomolecules in a potentiometric readout is
generally limited by the Debye screening of charges caused by charge screening from
counter ions. At elevated frequency levels (> 10 MHz) the principles are working in a
dielectric spectroscopy range, and the sensing is overcoming the limit of 1-2 nm [1, 2].
This is the fact for all frequency domain principles described above.
[1] S. Ingebrandt, Sensing beyond the limit, Nature Nanotech. 10, 734 (2015)
[2] C. Laborde et al., Real-time imaging of microparticles and living cells with CMOS nanocapacitor arrays, Nature Nanotech. 10, 791–795
(2015).
Biomolecules binding to silicon nanowires
What do we learn from the
combination of DC and AC?
DC signals:
Detection of charge
change and change of
local ionic environment
on surface
Electronic signals:
Specific biomolecule
binding – concentration of
analyte.
Binding of biomolecules to
the nanowire surface.
AC signals:
Detection of
capacitance change –
information about
density of molecule
layer and about
orientation
pH reference to cancel out
side effects.
Specific binding reaction on nanowires
Stern, E.; Vacic, A.; Li, C.; Ishikawa, F.N.; Zhou, C.W.; Reed, M.A.; Fahmy, T.M. A nanoelectronic enzyme-linked
immunosorbent assay for detection of proteins in physiological solutions. Small 2010, 6, 232–238.
42
05.02.2016
Outlook – Multiplexed biosensing
Summary
 Si nanowire arrays are fabricated in a wafer-scale process. They can be highly
integrated (up to 128x128 arrays) and the process is CMOS compatible
 Robust chips with reliable operation in liquid
 DNA and biomolecular detection experiments were successful
 Extracellular recordings are possible
Outlook
 Work out LODs for different assays (DNA hybridization, protein, immune)
 Model to explain the results
 Coupling with cells (mast cells, cardiac myocytes, tumor cell lines,
neuronal cells)
43
05.02.2016
Acknowledgements
FH Kaiserslautern
All members of AGBM and technical staff of FHKL
Financial support:
BMBF:
‘Nanowire Sensors’
‘Cancer Cell Chip’
‘Multiparametric Sensing’
DAAD:
PPP exchange grant – Germany/Hong Kong
Cooperation partners
FH Kaiserslautern
Profs. Karl-Herbert Schäfer, Monika Saumer,
Cornelia Keck
Research Center Jülich
Prof Dr. A. Offenhäusser
JL University Giessen
Prof. Dr. Martin Eickhoff
TU Kaiserslautern
Prof. Dr. Ing. Andreas König
University des Saarlands - Homburg
Prof. Dr. Markus Hoth
University of Applied Sciences Aachen
Prof. Dr. Ing. Michael J. Schöning
Universiteit Hasselt, Belgium
Prof. Dr. Patrick Wagner
The Chinese University Hong Kong
Prof. Dr. John A. Rudd
Prof. Dr. Chi-Kong Yeung
Italian Institute of Technology - Genua
Dr. Axel Blau
EU-ITN Marie Curie: Prosense
Industry: ‘Alternative Biosensor Principles’
Internal fh-funding:
- Nanotox
- Nanoimprint-Lithography
University of Applied Sciences Kaiserslautern
https://www.hs-kl.de/forschung/forschungsschwerpunkte/integrierte-miniaturisierte-systeme-ims/arbeitsgruppen/
44

Nanowire Sensing Systems - Nano-Tera

Transcription

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