Engineering, 2012, 5, 94-98
doi:10.4236/eng.2012.410B024 Published Online October 2012 (http://www.SciRP.org/journal/eng)
Electronic Detection of Escherichia coli O157︰H7 Using
Single-Walled Carbon Nanotubes Field-Effect Tr ansistor
Biosensor
Xiaoxian Zhang1, Dongwei Wang1, Danna Yang1, Sai Li1*, Zhiqiang Shen2
1School of Chemical Engineeri ng, Sichuan University, Chengdu, China
2Academy of Military Medical sciences, Chinese PLA Center for Disease Control & Prevention, Tianjin, China
Received 2012
ABSTRACT
Field effect tr ansisto rs (FET) based on Single-Walled Carbo n Nanotubes ( SWNTs) become the ho t topic in fields of nano-electronic,
clinical diagnostics, environmental testing etc. in recent years. In this paper, we reported a simple, scalable way to enrich semicon-
ducting SWNTs by using HNO3/H2SO4. Then carbon nanotube field-effect transistors (CNTFET) biosensor was fabricated with the
enrich ment SWNTs for E scherich ia coli O15 7︰H7 detect ion. The resp onse of each CN TFET was moni tored in real time before an d
after introduction of the Escherichia coli O157︰H7 at various con centration s. The results show that CNT-FET biosensors we fabri-
cated are sensitive t o chan ge of concen tration of so lution and response time is really short.
Keywords: Sin gle-walled Carbon Nanotubes (SWNTs); Field Effect Transistors; Biosensor; Escherichia Coli O157 ︰H7
1. Introduction
Biosensor plays an important role in many areas of environ-
mental monitoring, food analysis and life sciences. But tradi-
tional biological sensing techniques require multiple reagents,
signal amplification, and complex data analysis [1]. Because
SWNTs have outstanding properties such as possible biocom-
patibility, size compatibility and sensitivity towards minute
electrical perturbations [2], it has a great potential for biosens-
ing applications. Besides, CNTFET fabricated with semicon-
ducting SWNTs can operate at room temperature, reagentless
and l abel-free.
CNTFET has been used as biosensors to detect target bio-
molecules such as virus [3], bacterium [4,5], DNA [6] and en-
zymes [ 7]. Star et al. [8] h ave fabricated CNTFE T devices sen-
sitive to streptavidin by using individual biotin-functional- ized
CNTFET. Li et al. [9] repo rted the comple mentar y detectio n of
prostate-specific antigen (PSA) by using an anti-PSA monoc-
lonal antibody CNTFET. The mechanism of detection of bio-
molecules is still controversial. There are two major explains:
charge-transfer mechanism (electrostatic gating) [10] and
Schottky–Barrier modulation effect [11].
But there’s a hurdle to the widespread application of CNT-
FET. SWMTs synthesized are the mixture of semiconducting
SWNTs (sem-SWNTs) and metallic SWNTs (met-SWNTs).
The undesired met-SWNTs will hinder effective electronic
switching. Therefore, it’s essential to enrich or separate sem-
SWNTs from met-SWNTs. Many proposed methods were put
forward, such as d ielectr ophoresis [12], aromatic mol ecules [ 13]
and DNA wrapped [14]. But there’s not a scalable, nondestruc-
tive, iteratively repeatable and affordable sorting strategy
emerged [15].
In this paper, we separate metallic from semiconducting na-
notubes by using HNO3/H2SO4. Then we fabricate CNTFET
biosensor with treated SWNTs by applying AC dielectrophore-
sis [16] method. The CNTFET biosensor was functionalized
with linker molecule and the corresponding antibody. The
binding of the target E. coli O157︰H7 onto the receptor de-
tected by monitoring the electronic properties between source
and drain electrodes. We hope our work could contribute to fast
detection and comprehend of mechanism.
2. Materials and Methods
2.1. Materials
E. coli O157︰H7 antibody and E. coli O157︰H7 were sup-
plied by Academy of Military Medical sciences, Chinese PLA
Center for Disease Control & Prevention. 1-pyrenebutanoic
acid succinimidyl ester (PASE) was purchased from Sigma-
Aldrich. The reagents Dimethyl Formamide was purchased
from Kelong Chemical Company. All chemicals were used
without purification. CVD SWCNTs were produced by
Chengdu Organ ic Chemistry Research C enter.
2.2. Preparation of CNTFET Biosensor
The procedure of separate metallic from semiconducting nano-
tub es and fabricate C NTFET can be seen in our precious works
[17,18 ].
Immerse CNT channels in a 6 mM solution of PASE in di-
methyl formamide solution in room temperature overnight.
Then washed it with deionized water and blown dry with nitro-
gen. We can get a PASE-CNTFET. Secondly, the channels of
*Corresponding author.

YU. F. MIGAL ET AL.
95
P AS E -CNTFET were submerged in a 4.7 mg/mL solution of E.
coli O157︰H7 antibody in phosphate buffer solution (PBS)
overnight. The device surface was then thoroughly rinsed with
deionized water to remove the unreacted antibodies and dried
under a stream of nitrogen.
2.3. Detection of E. coli O157︰H7
Immerse ch ann els of ant ibody-CNTFET biosensors in solutions
of E. coli O157︰H7 at different concentrations of 8.2×102,
8.2×104, 8.2×106, 8.2×108 CFU/mL in PBS. The electrical
propert ies of the CNTFETs were measured in real time using a
semiconductor parameter analyzer. We also detected antibo-
dy-CNTFET which was not modified with PASE as a reference
to explore the impact of linker molecul e.
2.4. Characterization Techniques
The SWNTs pattern on the substrate was observed by a Scan-
ning Electron Microscopy (FE-SEM, Hitachi S4800) to eva-
luate the morphology of the samples. The CNTFETs have been
characterized at room temperature using an Agilent-4155C
semiconductor analyzer. Raman spectra were measured with a
HORIBA LabRAM HR Ramansc ope.
3. Results and Discussion
3.1. Enrichment of Sem-SWNTs
By contrast with Kataura plot, we can know the regions of
sem-SWNTs and met-SWNTs response relativity [19]. With
532nm laser, the peaks with wavenumbers 215-292cm-1 (0.8-
1.1nm) are due to met-SWNTs while those 135-215cm-1
(1.1-1.82) and 293-330cm-1 (0.74-0.8nm) are due to sem-
SWNTs. As shown in F igure 1( a), we can see an obvious in-
crease of sem-peaks around 160, 200 and 300cm-1 after
HNO3/H2SO4 treatments for 24h, whereas the met-peaks were
al mo s t ma i nt ai n same as pristin e S WNTs.
It may caused by charge transfer from SWNTs to NO2+. It’s
well known that mixing HNO3 and H2SO4 yields a high pro-
duction rate of NO2+ [20]. NO2+ as an electron less structure
can selectivity attack met-SWNTs because they have more
available charge density at the Fermi level than sem-SWNTs,
which induced t he selecti ve r emoval of met-S WN Ts .
Figure 1(b) demonstrates D and G band of SWNTs before
and after treatment. There’s an obvious drop in D band around
1450cm-1, which means a decreas e o f diso rder S WNTs, in other
words, treatment with HNO3/H2SO4 can purificate SWNTs as
well as enrich sem-S WN Ts.
3.2. Device Layout and Electrical Characterization of
CNTFET
Figure 2 shows the SEM image of electrode after SWNTs as-
sembly on Au electrodes. A lot of SWNTs bundles are arranged
between the sources and drain electrodes. F igure 3 reveals the
output characteristics of the CNTFET device fabricate with
enrichment of sem-SWNTs. From the transfer characteristics,
we can see that the drain current Id decreases with increasing
gate voltage Vg in the range between -10V and +10V. This
result indicated that CNT-FET showed p-type char acteristics in
the blank PB S buffer .
(a)
(b)
Figure 1. Raman spectra of pristine SWCNTs and HNO3/H2SO4(2:1)
treatment for 24h samples in RBM regoin (a) and G bands(b) un-
der 532 nm(data normalized with with peak at 260 cm−1 and G
band reference).
Figure 2. SEM image of CNTFET.

YU. F. MIGAL ET AL.
96
3.3. Detection of the E. coli O157︰H7 on CNTFET
Modified by Linker and Antibody
It’s a common way to functionalize SWNTs by using corres-
ponding receptor molecules as recognition layer to realize spe-
cific detecti on of bio molecul es. We detected E. col i O157︰H7
using CNTFET biosensors, in which CNTFET channels were
modified with E. coli O157︰H7 antibody. Linker bifunctional
molecule PASE can attach with antibody and SWNTs respec-
tively, playing a role as the bridge medium. The illustration for
fabrication of CNTFET immune biosensor was showed in Fig-
ure 4.
Resistance-volt a ge curves of CNTFET were demonstrated in
Figure 5 and Figure 6. Figure 5 shows that after modified
with PASE, the antibody-CNTFET biosensor causes a signifi-
cant decrease in the source-drain resistance contrast with the
pristine CNTFET. The significant change means E. coli O157
︰H7 could be captured by the antibody, then resistance change
follows. Figure 6 shows Resistance-voltage curve of CNTFET
after introduction of O157︰H7 into antibody-CNTFET with-
out PASE modified has slight change, which means CNTFET
without modified by linker could not attach effective antibody
to cap ture E. coli O157︰H7 .
Fig ure 3. Id -Vds curves of a CNTFET measured at room temper-
ature. (Vg =-10 to 10 V and steps are 5V).
Figure 4. Schematic illustration for fabrication of CNTFET
immu ne biosensor.
Figure 5. Resistance of Voltages of antibody-CNTFET after intro-
duction of O157︰H7 into antibody- CNTFET with PSE. a:
PASE-CNTFET, b: pristine CNTFET.
Figure 6. Resistance of Voltages of antibody-CNTFET after intro-
duction of O157︰H7 into antibody-CNTFET without
PASE.a:CNTFET witho u t PASE, b:pristine CNTFET .
After measuring of electrical properties of the CNTFE T bio-
sensors, we washed the device surface with deionized water
several times and dried under a stream of nitrogen. Then we
observed using SEM to confirm whether E. coli O157︰H7
were attached on channel of CNTFET or not.
Figure 7 shows SEM resu lts. CNTFET after P ASE modified
could attach E. coli O157︰H7 (Figure 7(a) ) whereas those
CNTFET without PASE could not catch the E. coli O157︰H7
(Figure 7(b)).
Combine R-V Curves and SEM results, we can get the con-
clusio n that the CENFET biosen sors cou ld realize t he effective
detection of E. coli O157︰H7. Only the CNTFET modified by
linker PASE and antibody have adsorption and response effect
on E. coli O157︰H7. The linker PASE plays an important role
to immobilize nanotubes and antibody. It’s due to the special
structure of PASE. It has two functional groups pyrene and
succinimidyl ester. Pyrene moiety could functionalize with the
SWNTs by π–π stacking and succinimidyl ester could react
with -NH2 in antibody to be a covalent bond.

YU. F. MIGAL ET AL.
97
(a)
(b)
Figure 7. SEM images of CNTFET after introduction of O157︰H7
(a) PASE-CNT FET (b ) CNTFET without PASE.
Time dependence of resistance of CNTFET after introduc-
tion of O157︰H7 at different concentration into antibody-
CNTFET was showed in Figure 8. Solution of concentrations
8.2×102, 8.2 ×104, 8.2×106, 8.2×108 CFU/mL of E. coli O157︰
H7 in PBS was dropped in channels of antibody-CNTFE T bio-
sensors. The electrical properties of the CNTFETs were meas-
ured in real time using a semiconductor parameter analyzer.
Resistan ce increases sharpl y to the maximum after dropped the
solution. Then it decreases rapidly in around 50s to a relative
stable level. Same trend occurred in different concentrations,
and maximum of resistance increased from 84kΩ to 110kΩ
along with the increase of concentration of E. coli O157︰H7
solution. The main reason is more E. coli O157︰H7 could be
captured by antibodies on CNTFET along with the increase of
concentration, then resistance between source and drain elec-
trode rises consequently.
The changes of resistance indicates the possibility of
CNTFET biosensor to detect E. coli O157︰H7, and concentra-
Figure 8. Time dependence of resistance of CNTFET after intro-
duction of O157︰H7 at different concentration into antibo-
dy-modifie d CNTFET.
tions as low as 8.2×102 CFU/mL can be readily detected with
response times of about 50s.
4. Conclusion
In this paper, we successfully fabricated CNTFET biosensor
with enriched semiconducting SWNTs by using a simple, scal-
able way of HNO3/H2SO4. Fr om SEM images and R-V cu rves,
we confirmed the effect of linker PASE. It can functionalize
with the SWNTs as well as the antibody. Then solutions of
concentrations 8.2×102, 8.2×104, 8.2×106, 8.2×108 CFU/mL of
E. coli O157︰H7 were detected with antibody-CNTFET bio-
sensor. There’s an obvious increase of resistance between
sour ce and drain with increas e of concen tration . We can draw a
conclusion that the CNTFET biosensors we fabricated are sen-
sitive to change of concentration of solution. The strategy can
be applied to general antibody-based detection schemes for
detecti ng concentratio n of target bi omolecules .
5. Acknowledgements
This research was supported by NSFC (20805033; 30901199),
SRF for ROCS, SEM (2008890-19-9).
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