Microfabricated Interdigitated Microelectrodes-Based ...
Transcript of Microfabricated Interdigitated Microelectrodes-Based ...
IBE Annual Meeting, March IBE Annual Meeting, March 66--9, 2008. Chapel Hill, NC9, 2008. Chapel Hill, NC 11
MicrofabricatedMicrofabricated InterdigitatedInterdigitatedMicroelectrodesMicroelectrodes--Based Based
Electrical/Electrochemical Impedance for Electrical/Electrochemical Impedance for Biological Detection Biological Detection
LijuLiju YangYang
BiomanufacturingBiomanufacturing Research Institute & Research Institute & Technology Enterprise (BRITE), Technology Enterprise (BRITE),
Department of Pharmaceutical Science, Department of Pharmaceutical Science, North Carolina Central University, Durham, NC North Carolina Central University, Durham, NC
2770727707
IBE Annual Meeting, March IBE Annual Meeting, March 66--9, 2008. Chapel Hill, NC9, 2008. Chapel Hill, NC 22
Impedance is a measurement of the ability of a circuit or electrical element to resist the flow of AC electrical current.
Solution
Impedance analyzer
Electrode
IBE Annual Meeting, March IBE Annual Meeting, March 66--9, 2008. Chapel Hill, NC9, 2008. Chapel Hill, NC 33
Conductive property change in the solution
Conductive property change at the interface of the electrodes
Impedance Change
1. Biological events occur in the solution.
2. Biological events occur at the interface of the electrodes
IBE Annual Meeting, March IBE Annual Meeting, March 66--9, 2008. Chapel Hill, NC9, 2008. Chapel Hill, NC 44
Interdigitated Array (IDA) Microelectrodes
Two-electrode systemMultiple electrode pairsProgressively shorter electrode distanceLarge electrode surface area
Low ohmic dropHigh signal-to-noise ratio Rapid attained steady stateImproved sensitivitySmall testing volumeReproducible fabrication
IBE Annual Meeting, March IBE Annual Meeting, March 66--9, 2008. Chapel Hill, NC9, 2008. Chapel Hill, NC 55
Chamber (50 µl)
Interdigitatedmicroelectrodes
Silicone rubber
Contact pads
Silicon substrate
AC signal generator
(a)
(b)
Van Gerwen P, et al. Sens. Actuators B, 1998; 49, 73-80
IBE Annual Meeting, March IBE Annual Meeting, March 66--9, 2008. Chapel Hill, NC9, 2008. Chapel Hill, NC 66
Impedance of biological cells
The cell membrane consists of a lipid bilayercontaining many proteins. It is highly insulating. The conductivity is around 10 -7 S/m.
The inside of a cell contains many dissolved charged molecules. The conductivity of the interior of a cell can be as high as 1 S/m.
IBE Annual Meeting, March IBE Annual Meeting, March 66--9, 2008. Chapel Hill, NC9, 2008. Chapel Hill, NC 77
(i) Making use of the metabolic activity of biological cells. This is represented by impedance microbiology, which is based on the measurements of the change in electric impedance in a medium or a reactant solution resulting from the bacterial metabolism.
Mechanisms for impedance detection of biological cells
(ii) Making use of the insulating properties of the cell membrane. –Label free biosensor.
(iii) Making use of the highly ionic cytoplasmof the cells. – cell lysis or ion release
IBE Annual Meeting, March IBE Annual Meeting, March 66--9, 2008. Chapel Hill, NC9, 2008. Chapel Hill, NC 88
180
200
220
240
260
280
0 2 4 6 8 10 12 14 16 18 20Time / h
Med
ium
Res
ista
nce
/ Ω No cell
9.3 x 103 CFU/ml 6.5 x 105 CFU/ml
7.6 x 101 CFU/ml
Conventional E at 1MHz
16
20
24
28
32
36
0 2 4 6 8 10 12 14 16 18
Growth time / h
Impe
danc
e / kΩ
0
104 103
102
101100105
IDA E at 10 Hz
0
2
4
6
8
10
12
14
16
1.0E+0 1.0E+1 1.0E+2 1.0E+3 1.0E+4 1.0E+5 1.0E+6
Concentration of Salmonella ( Log cfu/ml)
Det
ectio
n tim
e (h
)
Interdigitatedmicroelectrode
ConventionalElectrode
(i) Impedance detection based on bacterial metabolism
IBE Annual Meeting, March IBE Annual Meeting, March 66--9, 2008. Chapel Hill, NC9, 2008. Chapel Hill, NC 99
CdlCdl
Rsol
Cdi
Equivalent Circuit of the Microelectrode Systems
CdlCdl Rsol
Cdi
1.E+02
1.E+03
1.E+04
1.E+05
1.E+06
1.E+07
1.E-01 1.E+01 1.E+03 1.E+05 1.E+07Frequency / Hz
Impe
danc
e / Ω
MeasurementFitting
Cdl
RsolCdi
Cdl / nF Rs / Ω Cdi / pF
Before bacterial growth 397.2 ± 33.2 349.6 ± 16.0 45.3 ± 1.7After bacterial growth 528.2 ± 73.7 365.1 ± 25.2 44.6 ± 2.8
Change 33% 4.4% 1.5%
IBE Annual Meeting, March IBE Annual Meeting, March 66--9, 2008. Chapel Hill, NC9, 2008. Chapel Hill, NC 1010
(ii) Label-free impedance biosensor
Rs
Ret
Cdl
W
ic
if
iW
Ret
Cdl
-1200
-1000
-800
-600
-400
-200
00 500 1000 1500 2000 2500 3000
Real impedance / Ω
Imag
inar
y im
peda
nce
/ Ωb
e
d
ca
(a) antibodies
(b) 4.14x105
(c) 4.14x106
(d) 4.14x107
(e) 4.14x108
CFU/ml
IBE Annual Meeting, March IBE Annual Meeting, March 66--9, 2008. Chapel Hill, NC9, 2008. Chapel Hill, NC 1111
-180
-160
-140
-120
-100
-80
-60
-40
-20
00 100 200 300 400 500 600
Real Impedance (Ω )
Imag
inar
y Im
peda
nce
(Ω
)e-5um
e-10um
e-15um
e-20um
0.0E+0
1.0E+2
2.0E+2
3.0E+2
4.0E+2
5.0E+2
6.0E+2
1.0E+1 1.0E+2 1.0E+3 1.0E+4 1.0E+5
Frequency (Hz)
Impe
danc
e ( Ω
)
bare-5um
bare-10um
bare-15
bare-20um
10
210
410
610
810
1010
1210
10 20 30 40 50 60
Concentration of Fe(CN)6
Ret
5um
10um
15um
100
300
500
700
900
1100
1300
1500
1700
0 10 20 30 40 50 60
Cncentration of Fe(CN)6
impe
danc
e at
1 H
z
5um
10um
15um
IDA electrodes size effect
IBE Annual Meeting, March IBE Annual Meeting, March 66--9, 2008. Chapel Hill, NC9, 2008. Chapel Hill, NC 1212
Manipulating bacterial cells on sensor surfaces
No DEP
electrode
Negative DEP at 200 Hz and 3 Vpp
electrode
Positive DEP at 500 kHz and 3 Vpp
electrode
IBE Annual Meeting, March IBE Annual Meeting, March 66--9, 2008. Chapel Hill, NC9, 2008. Chapel Hill, NC 1313
0.6
0.8
1
1.2
1.4
1.6
1.8
17 19 21 23 25time (h)
cell
inde
x
500nM100nM20nM0.2nMcontrol
Oxytocin Dose-Dependent Response
0.00E+00
1.00E-01
2.00E-01
3.00E-01
4.00E-01
5.00E-01
6.00E-01
500 100 20 0.2 Control
Oxy Con (nM)
Cel
l Ind
ex
High throughput E-plates for label free cell-based drug screening
CHO Cells Over-Expressed OxytocinReceptor - Oxytocin Treatment
IBE Annual Meeting, March IBE Annual Meeting, March 66--9, 2008. Chapel Hill, NC9, 2008. Chapel Hill, NC 1414
(iii) Impedance detection based on ion release frombacterial cells
Glass substrate
Siliconerubber
Siliconerubber Impedance
analyzer
Interdigitated microelectrodesfor impedance measurements
Glass cover Cells in suspension
Electrode contact pads
1.0E+2
1.0E+3
1.0E+4
1.0E+5
1.0E+6
1.0E+0 1.0E+1 1.0E+2 1.0E+3 1.0E+4 1.0E+5
Frequency (Hz)
Impe
danc
e ( Ω
)
Measured dataFitting data
1.0E+2
1.0E+3
1.0E+4
1.0E+5
1.0E+6
1.0E+0 1.0E+1 1.0E+2 1.0E+3 1.0E+4 1.0E+5
Frequency (Hz)
Impe
danc
e ( Ω
)
Measured dataFitting data
Cdl RsCdl
CdlRs
Cdl
w wRet Ret
(A) In DI water (B) In PBS)
1.0E+2
1.0E+3
1.0E+4
1.0E+5
1.0E+6
1.0E+0 1.0E+1 1.0E+2 1.0E+3 1.0E+4 1.0E+5
Frequency (Hz)
Impe
danc
e ( Ω
)
Measured dataFitting data
1.0E+2
1.0E+3
1.0E+4
1.0E+5
1.0E+6
1.0E+0 1.0E+1 1.0E+2 1.0E+3 1.0E+4 1.0E+5
Frequency (Hz)
Impe
danc
e ( Ω
)
Measured dataFitting data
1.0E+2
1.0E+3
1.0E+4
1.0E+5
1.0E+6
1.0E+0 1.0E+1 1.0E+2 1.0E+3 1.0E+4 1.0E+5
Frequency (Hz)
Impe
danc
e ( Ω
)
Measured dataFitting data
1.0E+2
1.0E+3
1.0E+4
1.0E+5
1.0E+6
1.0E+0 1.0E+1 1.0E+2 1.0E+3 1.0E+4 1.0E+5
Frequency (Hz)
Impe
danc
e ( Ω
)
Measured dataFitting data
Cdl RsCdlCdl RsCdl
CdlRs
Cdl
w wRet Ret
CdlRs
Cdl
w wRet Ret
(A) In DI water (B) In PBS)
IBE Annual Meeting, March IBE Annual Meeting, March 66--9, 2008. Chapel Hill, NC9, 2008. Chapel Hill, NC 1515
1.0E+3
1.0E+4
1.0E+5
1.0E+0 1.0E+1 1.0E+2 1.0E+3 1.0E+4 1.0E+5
Frequency (Hz)
Impe
danc
e ( Ω
)w ater104 cells/ml105 cells/ml106 cells/ml107 cells/ml108 cells/ml109 cells/ml 1 kHz
(A)
1.0E+3
1.0E+4
1.0E+5
1.0E+0 1.0E+1 1.0E+2 1.0E+3 1.0E+4 1.0E+5
Frequency (Hz)
Impe
danc
e ( Ω
) PBS104 cells/ml105 cells/ml106 cells/ml107 cells/ml108 cells/ml109 cells/ml
(B)
109 cfu/ml
107 cfu/ml
108 cfu/ml
IBE Annual Meeting, March IBE Annual Meeting, March 66--9, 2008. Chapel Hill, NC9, 2008. Chapel Hill, NC 1616
1.0E+3
1.0E+4
1.0E+5
0 20 40 60 80 100Time (min)
Impe
danc
e ( Ω
)
109
103106
107105 104 102
108
water
y = -2.06 Log (x) + 5.23R2 = 0.98
0
5
10
15
20
25
30
35
40
1.0E+01.0E+21.0E+41.0E+61.0E+81.0E+10
Bacterial Concentration (cells/20µl)
Impe
danc
e at
1 k
Hz
(kΩ
)
Water
-25
-20
-15
-10
-5
0
5
0 5 10 15 20 25 30 35Time (h)
perc
enta
ge o
f im
peda
nce
chan
ge
(%)
PBS
PBS w ith cells
Water
Water w ith cells
IBE Annual Meeting, March IBE Annual Meeting, March 66--9, 2008. Chapel Hill, NC9, 2008. Chapel Hill, NC 1717
Acknowledgement Funding from Golden LEAF Foundation and the State of North Carolina through the Biomanufacturing Research Institute & Technology Enterprise (BRITE) Center for Excellence at North Carolina Central University.