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Capacitance immunosensors based on an array biotape
Chang Ming Li,*a Li Kun Pana and John H. T. Luongb
Received 19th April 2006, Accepted 23rd May 2006
First published as an Advance Article on the web 31st May 2006
DOI: 10.1039/b605651m
A new array immunosensing system with high-throughput has
been developed, based on the principle of a biotape that could
be used to make a biocassette recorder.
Immunosensing with high sensitivity and selectivity has been
developed for diversified applications in pharmaceutical, food,
clinical, forensic, and environmental monitoring.1–3 Although
reproducibility is always of utmost importance in any immuno-
assay format, the current trend and endeavour focus on the
development of label-free procedures for the detection of the
antigen–antibody binding event.2 Impedance measurements have
been used extensively in non-biological systems;4 however, this
technique is beginning to emerge in bio-analytical applications5–17
with its potential being reviewed by Bergveld.18 Instrumentation
has also been designed for accurately measuring minute changes
in impedance and conductance at regular time intervals. In
capacitance-based assays, the measurement is based on the
capacitance change owing to the formation of an antibody–
antigen complex on the electrode surface that affects a change
in the dielectric constant. Thus, the principle is simple,
straightforward, and requires no radioactive or fluorescent
labelling reagents. The technique is especially suitable for the
detection of high molecular-weight biomolecules such as DNA and
DNA fragments. To date, capacitance-based immunoassays have
been developed for important biomolecules such as hyaluronan-
binding proteins,19 human chorionic gonadotropin hormone
(hCG, a peptide hormone produced in pregnancy),20 transferrin,21
and anti-a-fetoprotein (an important tumor marker).3 The
detection limit is analyte-dependent, ranging from 0.5 pg mL21,
to 10 ng mL21. Very recently, a capacitance immunosensor based
on a self-assembled monolayer of thiourea on a gold electrode
has been reported with a detection limit of 10 pg mL21 for a
carcinoembryonic antigen standard.22 Conceptional capacitive
biosensing1 employs an electrode, submerged in an electrolyte
solution and the measurement is dependent upon the nature
and composition of the electrolyte buffer. The method can be
envisioned as a resemblance of a capacitor with the capacity to
store an electrostatic charge. Charged species and dipoles are
oriented at the electrode/solution interface, resulting in an electrical
double-layer. Primary screening in drug discovery, molecular
diagnostics and other bio-applications have fuelled immunoassay
a
School of Chemical & Biomedical Engineering, 50 Nanyang Avenue,
Singapore 639798. E-mail: ecmli@ntu.edu.sg; Fax: +65 67911161;
Tel: +65 67904485
b
National Research Council Canada, 6100 Royalmount Avenue,
Montreal, Quebec, Canada H4P 2R2.
E-mail: john.luong@cnrc-nrc.gc.ca; Fax: +1 514-496-6265;
Tel: +1 514-496-6175
788 | Analyst, 2006, 131, 788–790
www.rsc.
development towards the expansion of assays into multiplex and
high-throughput systems.
This paper describes a new sensing system with high-
throughput, based on the principle of a biotape to attain the
mobile electrical detection of the biomolecular recognition event.
The principle of capacitance measurement is based on the
parallel-plate capacitor theory. Two electrodes connected in
parallel, can be described as a capacitor with the resultant
capacitance of C = e0eA/d, where e0 is the vacuum permittivity,
e is the dielectric constant of the material between the plates,
d is the thickness, and A is the surface area. After coating with a
biocomposite dielectic layer, the capacitance will be affected,
depending upon its thickness and dielectric behavior (Fig. 1).
Addressable gold array electrodes on the tapes were made by
the PCB manufacturing process (Fig. 2). A homemade mani-
pulator was used for the impedance measurement. The prototype
was used to evaluate the concept and the system performance.
PCB is very soft with a thickness of 0.2 mm. The Au electrode on
the PCB is about 2 mm in diameter. Notice that the conventional
gold disk electrode is much larger than the gold array electrode. A
gold rod is positioned to engage the gold strip on the tape surface.
Two connecting leads from the two sensing electrodes are
connected to an impedance system.
Thioctic acid (11-mercapto-undecanoic acid), streptavidin, anti-
streptavidin, rabbit IgG, anti-rabbit IgG, goat IgG, anti-goat IgG
and bovine serum albumin (BSA) were purchased from Sigma-
Aldrich. Phosphate buffered saline (PBS) was used for buffer
preparation. Solutions were prepared from deionized water
(Ultrapure Water System Milli-Q Plus, Millipore).
The capacitance change of the electrode was evaluated in air
with ac impedance. The impedance measurement was performed
in the frequency ranging from 5000 Hz to 1.0 MHz under an
alternating voltage of 100 mV with the two-electrode measurement
method. The apparatus used for impedance measurement
consists of a Solartron Impedance Frequency Analyzer 1260 with
Impedance Interface 1294.
Fig. 1 (a) Schematic of capacitance measurement; (b) The corresponding
circuit model, where Cil and Cair are the capacitance of the insulating layer
and air, respectively.
This journal is ß The Royal Society of Chemistry 2006
Fig. 2 (a) A biotape prepared from a soft plastic sheet and (b) a
prototype system illustrates the biotape architecture with addressable gold
array electrodes deposited onto the flexible plastic tape surface. The
electrical contact to the individual electrodes is realized through pin
contact by applying pressure to a spring. Fig. 3 Plots of the capacitance difference (capacitance observed after
incubation at 25 uC with an antibody minus the baseline). (a) Anti-rabbit
Gold array electrodes on the surface of a tape shown in Fig. 2a IgG; (b) anti-rat IgG and (c) anti-streptavidin target. The PBS solution
were polished carefully with alumina slurries (1.0, 0.3, 0.05 mm), without anti-rabbit IgG, anti-rat IgG and anti-streptavidin is used as the
followed by sonication in distilled water and absolute ethanol. control.
Polishing of the disk electrodes on the tape was realized by rolling
the tape on sand paper. The clean electrodes were then immersed 10 pg mL21, respectively. Specificity was also performed using a
in a solution of 11-mercapto-undecanoic acid (linker) in absolute different pair of antigen and antibody. The result confirmed that
ethanol. After 6 h, the electrodes were rinsed with absolute ethanol the cross reaction rate of the different antibody target to the
and air-dried. The electrodes were then immerged in rabbit IgG, antigen probe was low, demonstrating excellent specificity of this
rat IgG, or streptavidin (100 mg mL21) in PBS solution. After 12 h labelless detection technology (Fig. 4).
of incubation, the electrodes were thoroughly rinsed with PBS, air-
dried and followed by the incubation with 3% BSA for 1 h. After
rinsing with PBS and air drying, the electrode impedance was
measured in air until a stable baseline was attained. After the
baseline reading, the electrodes were incubated with a solution
which consisted of anti-rabbit IgG, anti-rat IgG, or anti-
streptavidin for 2 h. The electrodes were then rinsed thoroughly
with PBS and air dried for 3–5 min, followed by the impedance
measurement for each electrode.
The imaginary part of impedance is in reverse proportion to the
frequency for a pure capacitor: Z0 = 1/(vC). Thus, the capacitance
C can be calculated by the slope of the straight line determined by
the variation of Z0 as a function of 1/v. The change in capacitance
for each electrode before and after incubation was thus obtained.
As shown in Fig. 3, before saturation, the capacitance variation
was linear with the logarithm of the antigen concentration and Fig. 4 Plots of the difference in capacitance of a baseline and that
anti-rabbit IgG, anti-rat IgG, and anti-streptavidin were deter- observed after incubation of the electrodes in different or corresponding
mined with a detection limit of 10 ng mL21, 10 ng mL21, and target solutions versus the type of probe and target molecules.

This journal is ß The Royal Society of Chemistry 2006 Analyst, 2006, 131, 788–790 | 789
 Notice that the immobilized antigen on the 11-mercapto-
undecanoic acid modified electrode was stable and after regenera-
tion, the response signal was established up to 30 times with an
RSD of y4%.
A simplified, sensitive and high throughput method, based on
the principle of the cassette recorder to fulfil the mobile electrical
detection of a protein has been developed. The experimental data
confirmed good sensitivity with high specificity of this novel
concept. The biotape concept can be used to make an x–y
addressable, high-throughput biochip device with simplicity of
manufacture process and great flexibility.
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This journal is ß The Royal Society of Chemistry 2006