Sensors and Actuators B 204 (2014) 211–217
Contents lists available at ScienceDirect
Sensors and Actuators B: Chemical
journal homepage: www.elsevier.com/locate/snb
ZnO nanorods as immobilization layers for interdigitated capacitive
immunosensors
P. Sanguino a,∗ , Tiago Monteiro a,b , S.R. Bhattacharyya c , C.J. Dias b , Rui Igreja b,∗∗ ,
Ricardo Franco a
a
REQUIMTE, Departamento de Química, Faculdade de Ciências e Tecnologia Universidade Nova de Lisboa 2829-516, Caparica, Portugal
CENIMAT/I3 N, Departamento de Ciência dos Materiais, Faculdade de Ciências e Tecnologia, FCT, Universidade Nova de Lisboa and CEMOP-UNINOVA
2829-516, Caparica, Portugal
c
Departamento de Física & ICEMS, Instituto Superior Técnico 1049-001, Lisbon, Portugal
b
a r t i c l e
i n f o
Article history:
Received 27 January 2014
Received in revised form 20 May 2014
Accepted 30 June 2014
Available online 1 August 2014
Keywords:
ZnO nanorods
Interdigitated electrodes
Capacitive immunosensor
Biosensor
Impedance spectroscopy
a b s t r a c t
ZnO nanorod structures were deposited on micrometer interdigitated Au electrodes to function as threedimensional matrixes for the immobilization of antibodies in a capacitive immunosensor format. As
a proof of concept, anti-horseradish peroxidase (anti-HRP) antibodies were immobilized on the ZnO
nanostructured surface by a crosslinking process. The ZnO nanorod layer allows distribution of antibodies across the entire region probed by the measuring electric field applied to the microelectrodes. This
is an alternative approach to the use of more expensive nanometer electrodes necessary in the detection of smaller layers of antibodies. The new micrometer interdigitated capacitive immunosensor was
able to discriminate between HRP antigen in buffer; a non-specific antigen in buffer; or buffer alone, as
proven by capacitance measurements. Maximum response of the sensor was achieved in the 5–6 kHz
frequency range, opening the possibility for a simplified single frequency detection system for direct
antigen detection in complex biological samples.
© 2014 Elsevier B.V. All rights reserved.
1. Introduction
Impedance spectroscopy is a powerful tool to detect physical,
chemical, or biochemical changes in a medium. In particular, in
the last 30 years, this technique has been extensively used as a
label free technique for the detection of biomolecular interactions
in the vicinity of metal or semiconductive electrode surfaces [1].
The application of this capability in the biosensors field was first
demonstrated in 1982 when Arwin et al. [2] showed that it was
possible to detect enzyme activity by the adsorption of protein on
the surface of an electrode. Later on, in 1986, Newman et al. [3]
reported the use of interdigitated electrodes (IDE) to detect capacitive changes from the specific binding of antibodies to antigens. This
affinity capacitive biosensor was built from two copper IDE with
a width and spacing of 50 ␮m which were covered with an insulating layer of parylene (1 ␮m thick) to prevent Faradaic currents
when measuring capacitance changes in a liquid environment. The
∗ Corresponding author. Tel.: +351 918 615 752.
∗∗ Corresponding author. Tel.: +351 212 948 562.
E-mail addresses: pesang@sapo.pt (P. Sanguino), rni@fct.unl.pt (R. Igreja).
http://dx.doi.org/10.1016/j.snb.2014.06.141
0925-4005/© 2014 Elsevier B.V. All rights reserved.
specificity of the biosensor was achieved by immobilization of
antigens on a SiO2 layer (0.3 ␮m thick) deposited on top of the
insulating polymer. The binding of the antibody to the immobilized
antigen created a change in the capacitance of the biosensor due
to changes on the permittivity of the small layer above the metal
IDE. The smaller size of these microelectrodes is an advantage over
macrosized traditional electrodes [4,5]. It allows for compactness,
portability, simplicity, higher sensitivity, and less volume of target
samples to be integrated in an immunosensor.
However, these early micrometer spaced electrodes affinity
biosensors were not able to create an appreciable impedance
variation upon binding of the antibody–antigen pair. This is not
surprising since, for micrometer IDE, the thickness of the layer
probed by the fringing electrical field above the electrodes (in the
micrometer range) is much larger than the sensing layer created
by the immobilized antigens or antibodies (typically 10–100 nm).
Therefore, this sensing layer, occupies just a fraction of the region
measured by the electric field which results in the small binding event signal being masked by the much larger signal from the
surrounding medium. This fact was confirmed by a theoretical calculation of the electric field between the IDE [6]. These authors
showed that all the current between the electrodes flows in a
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Fig. 1. (a) Schematic representation of the proposed immunosensor with the ZnO nanorod matrix layer for antibody immobilization. (b) Partial cross section of the
immunosensor. The ZnO nanorod layer is used to immobilize and distribute the antibody molecules across the region probed by the measuring electric field.
layer, thinner than the IDE spatial wavelength IDE , which in turn
is defined as 2 times the sum of electrode width W and electrode
gap G (IDE = 2(W + G)). For IDE with an equal gap and spacing of
10 ␮m this corresponds to a 40 ␮m layer. In fact, 95% of the current goes through a layer of 20 ␮m. More recently, Igreja and Dias
[7–9] used conformal mapping techniques to develop analytical
expressions for the capacitance of IDE. According to this model, the
capacitance of the IDE reaches a saturation value at a layer thickness
above the electrodes of IDE /2. For our IDE example with equal gap
and spacing of 10 ␮m this corresponds to 20 ␮m layer. Any physical, chemical, or biochemical change occuring beyond this distance
from the electrodes plane will have a negligible probability to be
measured as a capacitance variation by the interdigitated electrode
transducer.
In order to overcome the small thickness problem of the immobilization layer, and to increase sensitivity, IDE immunosensors
have been designed with nanometer electrode dimensions [6,10].
Ideal IDE values range from 20 to 200 nm. However, when compared to micrometer IDE, fabrication of nanometer electrodes
requires complex and expensive production methods and facilities. Nevertheless, most research groups have been developing
IDE immunosensors with dimensions of hundreds of nanometers
which is clearly a compromise between cost and sensitivity of the
biosensor [11–15].
Here, we investigated the use of ZnO nanorod structures
deposited by the hydrothermal method on Au micro-IDE (W and
G = 10 ␮m) as a means of immobilization of antibodies that will
function as probes to the target analyte of interest (Fig. 1a). The
3-dimensional matrix layer created by the ZnO nanorods allows
the distribution of the probe molecules across the region where
the fringing electric field penetrates. For our micro-IDE (W and
G = 10 ␮m) this region can spread to a distance of 20 ␮m above
the electrode plane (Fig. 1b). In this way, we expect these new
immunosensors to mostly detect capacitance changes due to the
binding affinity process and not to changes in the layer above the
small immobilization region as is the case in a conventional IDE
sensor where most of the probed region is air or liquid medium.
Owing to a number of interesting properties, ZnO is an ideal candidate to be used as the material for the matrix layer. It has high
mechanical strength, high thermal stability, and high oxidation
resistance in harsh environment. Its biocompatibility and nontoxicity makes it a strong candidate to be used in the bio and chemical
sensors field [16–20]. In addition, the fabrication of ZnO nanorod
structures by the hydrothermal route is simple, fast, inexpensive,
and can be performed at low temperature [21–23]. More recently,
ZnO nanorods were used for enzyme immobilization in glucose
sensors [24].
2. Materials and methods
2.1. Materials and reagents
The horseradish peroxidase (HRP) enzyme was from
Sigma–Aldrich, and used without further purification. Polyclonal
anti-HRP antibody was purchased from antibodies-online.com.
Tween 20, phosphate buffered saline (PBS) buffer, bovine serum
albumin (BSA), bicinchoninic acid (BCA), copper(II) sulfate,
2,2 -azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS),
(3-mercaptopropyl)trimethoxysilane
2,2,4-trimethylpentane,
(MPTMS), zinc nitrate hexahydrate (Zn(NO3 )2 ·6H2 O), and
sodium hydroxide were from Sigma–Aldrich. Sulfo-MBS (mmaleimidobenzoyl-N-hydoxysuccinimide ester) was from Pierce
Biotechnology. Hydrogen peroxide H2 O2 30% w/v and zinc acetate
dihydrate (Zn(CH3 COO)2 ·2H2 O) were from Panreac. Water was
Milli-Q quality. Amicon-ultra 500 (30 kDa) centrifugal filters were
from Millipore.
2.2. IDE fabrication
A double layer of Cr/Au (10/40 nm thickness) was deposited by
e-beam on a substrate of borosilicate glass. The thin Cr layer promotes adhesion of the gold layer. Micrometer IDE were patterned
according to standard photolithography and lift-off process. Fabricated microelectrodes had a width and gap of 10 ␮m which resulted
in a metallization ratio of 0.5 and a spatial wavelength IDE of
40 ␮m. The area covered by the microelectrodes was 0.7 cm2 .
2.3. ZnO nanorod deposition
Prior to deposition of the ZnO nanorods, the micro-IDE were
cleaned for 15 min in ethanol, acetone, DI water, and finally blown
dry with an Ar jet. The ZnO nanorod layer was deposited by the simple hydrothermal route in a two step process. In the first step, a seed
layer was deposited by spincoating a solution of zinc acetate dihydrate (Zn(CH3 COO)2 ·2H2 O) in ethanol (5 mM) for 30 s at 2000 rpm.
The coated microelectrodes were then dried with Ar jet, followed
by annealing in air at 250 ◦ C for 30 min. The annealing process promotes the thermal decomposition of the zinc acetate to ZnO seeds
according to the following equation:
Zn(CH3 COO)2 → ZnO + CO2 + (CH3 )2 CO
(1)
For each interdigitated microelectrode, this step was repeated
twice in order to increase the density of the nucleation layer.
For the hydrothermal growth, the ZnO seeded microelectrodes
were suspended in a 0.01 M aqueous solution of zinc nitrate
P. Sanguino et al. / Sensors and Actuators B 204 (2014) 211–217
213
hexahydrate (Zn(NO3 )2 ·6H2 O) and 0.35 M NaOH. Separate solutions of zinc nitrate hexahydrate and NaOH were prepared in
deionized water (Mili-Q). The solutions were then mixed in a
glass beaker containing the suspended microelectrodes, by slowly
adding the NaOH solution to the zinc nitrate hexahydrate solution.
After vigorously stirring (700 rpm) the solution for 2 h at 25 ◦ C, the
temperature was increased to 80 ◦ C and the stirring was reduced
to 300 rpm. After 6 h of deposition, the ZnO coated microelectrodes
(micro-IDEZnO ) were dried with an Ar jet followed by 30 min at
90 ◦ C for drying. Crystal growth orientation of ZnO nanorods was
analyzed by X-ray diffraction in a –2 configuration equipped with
a Cu source.
2.4. Antibody immobilization
Attachment of the antibody probes to the ZnO nanorod
structures was by cross-linking with Sulfo-MBS. This heterobifunctional crosslinker contains N-Hydroxysuccinimide (NHS) ester and
maleimide groups that allow covalent conjugation with aminoacid
side chains presenting amine and sulfhydryl groups. The antibody immobilization process is divided in 3 stages: silanization,
conjugation and crosslinking. In the silanization step (Fig. 2(a)),
the micro-IDEZnO transducers were submerged in a 2% (v/v) solution of MPTMS in 2,2,4-trimethylpentane for 1 h. To remove the
unbounded MPTMS, the silanized transducers were then washed
in the solvent and finally dried with an Ar jet. Attachment of the
MPTMS molecules to the ZnO surface has been reported to be predominantly through the silane groups with the sulfhydryl groups
molecularly oriented away from the surface [25]. Therefore, this
stage makes sulfhydryl ( SH) groups available at the surface of the
nanorods for further linking to maleimide groups of the crosslinker
Sulfo-MBS. In the antibody conjugation stage (Fig. 2(b)), a PBS solution of anti-HRP antibody (0.5 mg/ml) and Sulfo-MBS crosslinker
(13.8 ␮g/ml) was incubated for 30 min at room temperature. The
moderate 10-fold molar excess of crosslinker over antibody was
selected in order to avoid deleterious effects to the antibody structure.
Covalent conjugation is possible by the attachment of the NHS
ester groups of Sulfo-MBS to the amine groups of the antibody antiHRP. The unbounded crosslinker, was removed by ultrafiltration
with an Amicon-Ultra 500 (30 kDa) and centrifugation at 7500 × g,
15 min at 4 ◦ C. Finally, in the crosslinking stage (Fig. 2(c)), 50 ␮l
of the conjugated mixture were spread over the silanized IDEZnO
transducer and left to incubate for 3 h at room temperature. During
this process, the maleimide groups of the conjugate S-MBS/antiHRP bind to the sulphydryl groups present on the silanized surface
of the ZnO. The prepared sensors were washed in PBS solution and
dried with Ar jet. Following the immobilization of antibody, and
in order to prevent non-specific binding, the sensors were covered
with 50 ␮l of 5% (w/v) BSA in PBS with 0.05% (v/v) Tween®-20 and
left to incubate for 2 h. Finally, the mounted sensor set was washed
3 times for 5 min in plenty of fresh PBS.
2.5. Antigen sensor testing
To test the IDEZnO immunosensor, we applied 50 ␮l of PBS, HRP
in PBS (6.7 ␮M), and an unrelated and non-specific antigen (Plasmodium falciparum Heat shock protein 70 – PfHsp70) (6.7 ␮M in
PBS) on 3 identical immunosensors that were fabricated simultaneously. The testing solution was left on the sensors for 1 h at room
temperature and then washed with PBS. In the following, SHRP , SPBS ,
and SPfHsp70 refer to sensors tested with HRP, PBS, and PfHsp70 solutions, respectively. On these tests, HRP is the specific antigen target
and PfHsp70 the non-specific antigen.
Fig. 2. Antibody immobilization on ZnO nanorods with S-MBS crosslinker. (a)
Silanazation of the ZnO nanorod surface. (b) Conjugation of anti-HRP antibody with
the crosslinking moiety. (c) Crosslinking of the conjugate to the silanized nanorods.
2.6. Measurements
Capacitance (C) and loss tangent (tan ı) measurements were
made with an Agilent 4294A precision impedance analyzer in the
range of 40 Hz to 110 MHz. Measurements were performed after
each fabrication and testing stage. To eliminate the influence of
water (either in the liquid or vapor forms), sensors were dried for
30 min under vacuum. After reaching the base pressure of the system (1E−2 mbar), C values were recorded for all spectra. For each
sensor, 3 measurements were taken where the electrical contacting
probes were repositioned each time.
2.7. Horseradish peroxidase (HRP) enzymatic assay
The presence of antigen (due to specific binding to the respective antibody) in the tested sensors was assessed by an enzymatic
assay of horseradish peroxidase (HRP) using 2,2 -azino-bis(3ethylbenzothiazoline-6-sulphonic acid) (ABTS) as substrate. The
three sensors, SHRP , SPBS , and SPfHsp70 , were placed inside a solution
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Fig. 3. Deposition of ZnO nanorods on Au micro-IDE. Density of ZnO nanorods clusters over the micrometer electrodes for deposition times of (a) 3, (b) 5, and (c) 7 h. Stability
of ZnO nanorods: (d) before washing and (e) after washing twice with PBS.
of ABTS + H2 O2 according to the Sigma–Aldrich enzymatic assay of
peroxidase [26]. In brief, the amount of HRP enzyme present in the
sensors is proportional to the amount of oxidized ABTS which can
be determined by measuring the absorbance at 424 nm and visually confirmed by the development of a green color in solution. This
color change was evaluated 3 min after submerging the sensors in
the assay solution.
3. Results and discussion
3.1. ZnO nanorod layer
The first step in the fabrication of the proposed immunosensor was the hydrothermal deposition of the ZnO nanostructures on
top of the Au micro-IDE. Photoluminescence and photoconductivity measurements of nanorod structures obtained with this method
have been previously reported [27]. Ideally, the 3-dimensional
layer should have a high density of ZnO structures with sizes capable of including them well inside the measuring region of the
probing electric field. The duration of the hydrothermal process
is an important parameter for determining the size and coverage
of ZnO structures deposited on the Au microelectrodes. In order
to establish the deposition time to be used in the fabrication of
the sensors, ZnO nanostructures were grown on Au micro-IDE for
three different deposition times: 3, 5, and 7 h (Fig. 3). From Fig. 3(a)
we can conclude that a deposition time of 3 h did not produce an
acceptable coverage of the electrodes. On the other hand, for 5 h
(Fig. 3(b)) and 7 h (Fig. 3(c)) of deposition, a much higher density of
nanorod structures was observed.
Apart from ZnO nanostructures density and size, the stability
of ZnO nanostructures is paramount in the fabrication and testing of the proposed immunosensor. To determine this parameter, a
micro-IDEZnO was washed with PBS to simulate the fabrication and
testing procedures. After drying with an Ar jet, the washing procedure was repeated. Fig. 3(d) and (e) shows optical microscopy
images of the micro-IDEZnO before and after washing twice with
PBS, respectively, showing that ZnO nanorod structures remained
attached to the surface of the microelectrode transducer. It can
therefore be concluded that these 3-dimensional structures are
optimal anchoring substrates for antibody probe immobilization
on the immunosensor.
In order to shorten sensor fabrication time as much as possible, not compromising the quality of the obtained sensors, we
have decided for a deposition time of 6 h. In these conditions,
we obtained ZnO nanorods with approximate lengths of 5 ␮m
and widths of 100 nm (Fig. 4(a)). The X-ray diffraction pattern
of Fig. 4(b) shows that the deposited ZnO nanorods exhibit the
wurtzite hexagonal crystal structure with preferential growth in
Fig. 4. (a) SEM micrograph of the ZnO nanostuctured matrix layer. (b) –2 X-ray diffraction pattern for the deposited ZnO nanorods. The first peak, located at about 34.5◦ ,
can be assigned to the (0 0 2) crystallographic plane family in hexagonal ZnO.
P. Sanguino et al. / Sensors and Actuators B 204 (2014) 211–217
Fig. 5. Evolution of the capacitance (at 10 kHz; log scale) for the three sensors during
the various fabrication steps. The deposition of ZnO nanorods increases the capacitance of the micro-IDE by two orders of magnitude. During fabrication, changes
in capacitance are identical for all the sensors. Sensors SHRP , SPf Hsp70 , and SPBS were
tested with specific antigen, non-specific antigen and buffer, respectively.
the (0 0 2) direction. This fact can be attributed to the low surface
energy of the (0 0 2) crystal plane [28]. The XRD peak at 38.4 was
assigned to Zn (1 0 0) according to the Joint Committee on Powder
Diffraction Standards (JCPDS) card (no. 040831).
3.2. Sensor fabrication
215
Fig. 6. Confirmation of the presence of HRP in the immunosensor by an enzymatic
assay of horseradish peroxidase (HRP) using ABTS as substrate. The sensor tested
with HRP solution was able to oxidize ABTS as indicated by the green color change.
Sensors tested with PBS and PfHsp70 were not able to promote this color change,
confirming HRP absence.
The response of the sensors to the testing solutions was determined by calculating the capacitance and phase shift variation from
the measured capacitance and loss tangent spectra. Fig. 7 presents
these variations when sensors are tested with the respective assay
(HRP; specific antigen), or control (PfHsp70; non-specific antigen),
and PBS solutions.
When the immunosensor was tested with the solution containing the specific antigen (anti-HRP), a decrease in the capacitance
Having determined the conditions for the hydrothermal deposition of ZnO nanostructures, three sensors (SHRP , SPBS , and SPfHsp70 )
were fabricated simultaneously as described in the materials and
methods section. Briefly, the process can be divided into four
stages: (1) deposition of ZnO nanostructures on the surface of
the micro-IDE; (2) functionalization of the ZnO nanostructured
electrodes with the organosilane MPTMS; (3) cross-linking of the
SMBS/anti-HRP conjugate to the MPTMS treated electrodes; (4)
blocking of the sensors with BSA, to prevent nonspecific binding.
The capacitance of the three immunosensors was recorded after
each fabrication step. As presented in Fig. 5, after deposition of the
ZnO nanostructures (first step of the fabrication process), the capacitance of the micro-IDE increased by two orders of magnitude. This
capacitance increase can be explained by the increased surface area
provided by the ZnO nanostructures. After the silanization step,
the capacitance increases again for all the sensors by almost one
order of magnitude. However, a slight decrease of the capacitance
takes place after the two following fabrication steps, namely, antibody crosslinking and BSA blocking. It is important here to notice
that shifts in the capacitance after each fabrication step, are always
in the same direction for all three sensors. This identical behavior
attests the robustness of the fabrication procedure of the proposed
immunosensors.
3.3. Sensor testing
Finalized sensors, with a nominal capacitance of 10 nF (Fig. 5),
were tested with the respective assay solutions: PBS only for the
SPBS sensor; anti-HRP in PBS, for the SHRP sensor; and PfHsp70 in
PBS, for the SPfHsp70 sensor. The presence of the antigen in the SHRP
sensor was confirmed by an enzymatic assay of horseradish peroxidase (HRP) (see Section 2). The green color change, corresponding
to ABTS oxidation and promoted by the presence of HRP in the sensors, was only seen in the sensor tested with HRP solution (Fig. 6).
The sensors SPBS and SPfHsp70 that did not contain HRP, did not promote the oxidation of the ABTS solution and consequently, no green
color was observed.
Fig. 7. (a) Sensors capacitance (real part) and (b) phase shift variation spectra of the
IDEZnO immunosensor when tested with an HRP antigen solution, PfHsp70 nonspecific antigen solution, and PBS buffer alone.
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was observed for all tested frequencies below 1 MHz (Fig. 7(a)). On
the other hand, testing the sensor with the non-specific solutions
(PBS only or PfHsp70 in PBS) resulted in the increase of the capacitance in the same frequency range. This opposite response confirms
that the proposed immunosensor is able to discriminate between
solutions containing the specific target (anti-HRP) and solutions
that do not contain this HRP antibody-specific antigen. The buffer
solution (PBS) in which the specific and non-specific antigens
are diluted will affect the immunosensor response, increasing the
capacitance. This is probably due to accumulation of Na+ and Cl−
ions in the sensor matrix which in turn increases its permittivity. This could explain the positive variation of the immunosensor
capacitance when tested with PBS and non-specific antigen solutions. After antigen-antibody bonding the medium permittivity will
decrease which is attributed to a decrease in antibody polarization
when bonded to the specific antigen. The same opposite response
between controls and specific antigen is also obtained when plotting the phase shift variation below 8 kHz (Fig. 7(b)), but in this
case, the response of the sensor to the specific antigen solution is
positive, while the response to the non-specific solutions is negative.
It is interesting to notice that the variation of the capacitance
(Fig. 7(a)), when testing the immunosensor with the specific antigen (HRP), decreases for frequencies above 6 kHz. Based on the
physical model described by Pethig and Kell [29], Gebbert et al.
[30] calculated that the maximum excitation frequency at which
an immobilized antibody–antigen complex (in a water medium)
responds to changes in the electric field, is around 6 kHz. This is
consistent with our data. Although our immunosensors are in a
“dry” state during measurements, the water vapor present in the
low vacuum chamber (10−2 mbar) is probably enough to keep a
water envelope around the antibody–antigen complex, therefore
validating the calculations for our case.
The maximum variation of the capacitance in the 5–6 kHz frequency range allows for simplified single frequency probing in
future practical applications.
4. Conclusions
For the first time, we have used ZnO nanostructures as a
sensitive layer coupled with interdigitated microelectrode transducers in an affinity immunosensor format. The addition of a
3-dimensional matrix, on top of the interdigitated microelectrodes, allowed distribution of antibody probes across the region
probed by the fringing electric field. This setup opens the opportunity for the use of interdigitated electrodes with micrometer
dimensions (10 ␮m), an important advantage over nanometer
interdigitated sensors for which more complex and expensive fabrication procedures are required. The new IDEZnO immunosensors
were able to identify buffered aqueous solutions containing the
target antigen (HRP), responding with a negative capacitance variation. Conversely, solutions without the specific antigen, produced
an opposite variation in the capacitance of the immunosensors.
The IDEZnO immunosensor response presented a maximum at the
excitation frequency of 6 kHz which foresees the future sensor
instrumentation particularly easy for practical applications.
Acknowledgments
This work was supported by Fundação para a Ciência e
a Tecnologia, Portugal (Grants PEstC/EQB/LA0006/2013 to
PS and RF; PTDC/CTM-NAN/112241/2009 to RF; post-doc
grant SFRH/BPD/70803/2010 to PS; post-doc grant SFRH/BPD/
66773/2009 to SRB; and the COMPETE Program). We thank
Mafalda Costa for help in optical microscopy.
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Biographies
Pedro Sanguino received his degree in Physics Engineering from Instituto Superior
Técnico, IST, Lisbon, Portugal, in 1999 and the Ph.D. degree in Technological Physics
Engineering from the same institution in 2005. Since 2011 he is a Postdoc in the
Chemistry Department of Faculdade de Ciências e Tecnologia from the Universidade
Nova de Lisboa (FCT-UNL). Dr. Sanguino has a solid background in the production
and characterization of thin films and is currently working in the biosensors field.
Tiago Monteiro received the degree in Molecular and Cellular Biology from Universidade Nova de Lisboa (UNL) in 2011. He is currently pursuing his M.Sc. Degree in
Biotechnology at the same institution.
Soumya Bhattacharyya was born in Kolkata/India in 1981. He completed his studies
of Physics in 2004 and began his PhD in 2005 with a CSIR fellowship at the Jadavdur
University. His thesis topic was “Synthesis and charaterization of Gallium Nitride
films by sputtering technique”. He completed his PhD in 2009 and worked as a Postdoc fellow (FCT grant) at the Instituto Superior Técnico in Lisbon. He is currently
an assistant professor at the physics department of Suri Vidyasagar College in India.
His current research interests are wide band gap semiconductors and their device
applications.
C.J. Dias was born in Angola in 1959. He graduated in Engineering Physics from
Universidade Nova de Lisboa (UNL) in 1982 and joined the Physics Department of
the same University in 1985. He received the Ph.D. from University of North Wales
(Bangor) in 1994 working on ferroelectric polymer–ceramic composites. Since then
Prof. Dias has been auxiliary professor at UNL and is currently working in the Materials Science Department and is a full member of CENIMAT/I3N research associated
laboratory in Nanostructures, Nanomodelation and Nanofabrication. His research
P. Sanguino et al. / Sensors and Actuators B 204 (2014) 211–217
interests include characterization of dielectrics and their use in various applications such as sensors and actuators, acoustics and polymer materials for energy
applications.
Rui Igreja was born in Lisbon, Portugal in 1966. He graduated in Engineering
Physics from Universidade Nova de Lisboa (UNL) in 1992 and joined the Physics
Department of the same University in 1993. He obtained the M.Sc. degree in
instrumentation, industrial maintenance and quality in UNL in 1998 and the
Ph.D. in materials engineering (microelectronics and optoelectronics) (2006) in
UNL. Prof. Igreja is currently an auxiliary professor at Materials Science Department at UNL and full member of CENIMAT/I3N research associated laboratory in
217
nanostructures, nanomodelation and nanofabrication. His research interests include
solid state physics, dielectrics and chemical and biological sensors.
Ricardo Franco (b. 1966) holds a degree in Applied Chemistry and Biotechnology
(1989) and a Ph.D. in Bioinorganic Chemistry (1995), both by Universidade Nova
de Lisboa, Portugal. Currently, he is an assistant professor of Biochemistry at the
Faculdade de Ciências e Tecnologia, Universidade Nova de Lisboa, Portugal. Prof.
Franco’s research activities focus on antibodies, protein and DNA interactions with
nanoparticles and nanostructures of noble metals using spectroscopic techniques,
for the development of nanobiosensors.