Biosensors & Bioelectronics 17 (2002) 345 359

www.elsevier.com/locate/bios

Review

Application of conducting polymers to biosensors

Manju Gerard a, Asha Chaubey b,*, B.D. Malhotra b
b

a
Chemistry Department, Allahabad Agricultural Institute (Deemed Uni6ersity), Allahabad, India
Biomolecular Electronics and Conducting Polymer Research Group, National Physical Laboratory, Dr. K.S. Krishnan Marg, New Delhi 110
012, India

Received 14 August 2001; accepted 31 October 2001

Abstract
Recently, conducting polymers have attracted much interest in the development of biosensors. The electrically conducting
polymers are known to possess numerous features, which allow them to act as excellent materials for immobilization of
biomolecules and rapid electron transfer for the fabrication of efficient biosensors. In the present review an attempt has been made
to describe the salient features of conducting polymers and their wide applications in health care, food industries, environmental
monitoring etc. 2002 Elsevier Science B.V. All rights reserved.
Keywords: Conduction polymer; Biosensor; Immobilization; Electron transfer; Tranducer

Contents
1. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
1.1. Conducting polymers. . . . . . . . . . . . . . . . . . . . . . . .
1.2. Historical background of electronically conducting polymers.
1.3. Conduction mechanism . . . . . . . . . . . . . . . . . . . . . .
1.4. Synthesis of conducting polymers . . . . . . . . . . . . . . . .
1.5. Applications of conducting polymers . . . . . . . . . . . . . .
2. Biosensors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
2.1. Transducer . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
2.2. Biocomponents . . . . . . . . . . . . . . . . . . . . . . . . . . .
3. Importance of conducting polymers to biosensors . . . . . . . . . .
4. Immobilization of enzymes on conducting polymers . . . . . . . .
5. Types of biosensors . . . . . . . . . . . . . . . . . . . . . . . . . . .
5.1. Amperometric biosensors . . . . . . . . . . . . . . . . . . . . .
5.2. Potentiometric biosensors . . . . . . . . . . . . . . . . . . . . .
5.3. Conductometric biosensors . . . . . . . . . . . . . . . . . . . .
5.4. Optical biosensors . . . . . . . . . . . . . . . . . . . . . . . . .
5.5. Calorimetric biosensors . . . . . . . . . . . . . . . . . . . . . .
5.6. Peizoelectric biosensors . . . . . . . . . . . . . . . . . . . . . .
6. Applications of conducting polymers . . . . . . . . . . . . . . . . .
6.1. Biosensors for health care . . . . . . . . . . . . . . . . . . . . .
6.1.1. Glucose biosensors . . . . . . . . . . . . . . . . . . . . .
6.1.2. Urea biosensors . . . . . . . . . . . . . . . . . . . . . . .
6.1.3. Lactate biosensors . . . . . . . . . . . . . . . . . . . . .

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

* Corresponding author. Fax: + 91-11-585-2678.

E-mail address: achaubey@csnpl.ren.nic.in (A. Chaubey).
0956-5663/02/$ - see front matter 2002 Elsevier Science B.V. All rights reserved.
PII: S 0 9 5 6 - 5 6 6 3 ( 0 1 ) 0 0 3 1 2 - 8

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.

346
346
346
347
347
347
348
348
349
349
350
351
351
351
352
353
353
353
353
353
353
353
354

346

M. Gerard et al. / Biosensors & Bioelectronics 17 (2002) 345359

6.1.4. Cholesterol biosensors . . . . . . .

6.2. Immunosensors . . . . . . . . . . . . . . .
6.3. DNA Biosensors . . . . . . . . . . . . . .
6.4. Biosensors for environmental monitoring
6.5. Biosensors for food industry . . . . . . .
7. Conclusion . . . . . . . . . . . . . . . . . . . .
Acknowledgements. . . . . . . . . . . . . . . .
References. . . . . . . . . . . . . . . . . . . . .

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

1. Introduction
Polymers are being discarded for their traditional
roles as electric insulators to literally take charge as
conductors with a range of novel applications. Scientists from many disciplines are now combining expertise
to study organic solids that exhibit remarkable conducting properties. A large number of organic compounds, which effectively transport charge are roughly
divided into three groups i.e. charge transfer complexes/
ion radical salts, organometallic species and conjugated
organic polymers. A new class of polymers known as
intrinsically conducting polymers or electroactive conjugated polymers has recently emerged. Such materials
exhibit interesting electrical and optical properties previously found only in inorganic systems. Electronically
conducting polymers differ from all the familiar inorganic crystalline semiconductors e.g. silicon in two important features that polymers are molecular in nature
and lack long range order (Duke and Schein, 1980). A
key requirement for a polymer to become intrinsically
electrically conducting is that there should be an overlap of molecular orbitals to allow the formation of
delocalized molecular wave function. Besides this,
molecular orbitals must be partially filled so that there
is a free movement of electrons throughout the lattice
(Bloor and Movaghar, 1983).

1.1. Conducting polymers

Conducting polymers contain (p-electron backbone
responsible for their unusual electronic properties such
as electrical conductivity, low energy optical transitions,
low ionization potential and high electron affinity. This
extended (p-conjugated system of the conducting polymers have single and double bonds alternating along
the polymer chain. The higher values of the electrical
conductivity obtained in such organic polymers have
led to the name synthetic metals. Many applications
of conducting polymers including analytical chemistry
and biosensing devices have been reviewed by various
researchers (Trojanowicz and vel Krawczyk, 1995; Situmorang et al., 1998; Schuhmann, 1995; Wring and
Hart, 1992; Guiseppi-Elie et al., 1997). They have
widened the possibility of modification of surface of
conventional electrodes providing new and interesting
properties. They were applied in electrocatalysis, mem-

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

.
.
.
.
.
.
.
.

354
354
354
354
355
355
355
355

brane separations and chromatography. They also create new technological possibilities in design of chemical
and biochemical sensors (Trojanowicz and vel
Krawczyk, 1995; Bidan, 1992; Bartlett and Cooper,
1993).

1.2. Historical background of electronically conducting

polymers
Research on conducting polymers intensified soon
after the discovery of poly(sulphur nitride) [(SN)x] in
1975 which becomes superconducting at low temperatures (Greene et al., 1975). Although, conducting polymer complexes in the form of tetracyano and
tetraoxalato-platinates, the Krogman salts charge transfer complexes (Minot and Peristein, 1971) had been
known earlier, the significance lies in the rediscovery of
PA in 1977 (initially discovered by Shirakawa et al.,
1977 using a Ziegler Natta type polymerization catalyst) by MacDiarmid and Heeger, University of Pennsylvania. They were able to enhance the electrical
conductivity of PA (10 9 S cm 1) by several orders i.e.
105 S cm 1 by simple doping with oxidizing agents e.g.
I2, AsF5, NOPF6 (p-doping) or reducing agents (n-doping) e.g. sodium napthalide. This has generated renewed interest of the scientific community towards the
study and discovery of new conducting polymeric
systems.
Poly-paraphenylene was synthesized by Ivory et al.
(1979). It forms highly conducting charge transfer complexes with both n and p type dopants. Doping with
AsF5 increases its conductivity to its values from 10 5
to 500 cm 1. Theoretical models and electron spin
resonance measurements indicate that the charge transport in PPP is a polaron/bipolaron. PPS was the first
non-rigid, but not fully carbon backbone linked conducting polymer. Its discovery was particularly exciting,
since its property of solution processability opened the
door for potentially obtaining commercially viable conducting plastics (Rabolt et al., 1980).
Amongst polyheterocyclines, polypyrrole (PPY) has
been investigated the most. The electrochemical oxidation of pyrrole in aqueous H2SO4 can be carried out on
platinum electrode. The product is a conducting polymer known as Pyrrole Black Kanazawa et al. (1979)
produced coherent films of PPY with a conductivity of
100 Scm 1 and exhibited excellent air stability. But the

M. Gerard et al. / Biosensors & Bioelectronics 17 (2002) 345359

main hindrance of its processability is in its insolubility

in any organic solvents.
PTH shows remarkable stability of both oxidized
(p-doped) conducting form and its neutral (undoped)
insulating form in both air and water. It shows high
doping level upto 50% which may be attributed to its
partially crystalline nature that has been confirmed by
X-ray photoelectron spectroscopy studies (Tourillon
and Garnier, 1983). Many other conducting polymers
such as polyfuran, polyindole, polycarbazole, polyaniline etc. have also been synthesized. Structures of some
typical conducting polymers have been shown in Fig. 1

1.3. Conduction mechanism

The mechanism of conduction in such polymers is
very complex since such a material exhibits conductivity
across a range of about fifteen orders of magnitude and
many involve different mechanisms within different
regimes. Conducting polymers show enhanced electrical
conductivity by several orders of magnitude of doping.
The concept of solitons, polarons and bipolarons has
been used to explain the electronic phenomena in these
systems (Heeger, 1986). Conductivity in conducting

347

polymers is influenced by a variety of factors including

polaron length, the conjugation length, the overall
chain length and by the charge transfer to adjacent
molecules (Kroschwitz, 1988). These are explained by
large number of models based on intersoliton hopping,
hopping between localized states assisted by lattice
vibrations, intra-chain hopping of bipolarons, variable
range hopping in 3-dimensions and charging energy
limited tunneling between conducting domains.

1.4. Synthesis of conducting polymers

Various methods are available for the synthesis of
conducting polymers. However, the most widely used
technique is the oxidative coupling involving the oxidation of monomers to form a cation radical followed by
coupling to form di-cations and the repetition leads to
the polymer. Electrochemical synthesis is rapidly becoming the preferred general method for preparing
electrically conducting polymers because of its simplicity and reproducibility. The advantage of electrochemical polymerization is that the reactions can be carried
out at room temperature. By varying either the potential or current with time the thickness of the film can be
controlled.
Electrochemical polymerization of conducting polymers is generally employed by: (1) constant current or
galvanostatic; (2) constant potential or potentiostatic;
(3) potential scanning/cycling or sweeping methods.
Standard electrochemical technique which employs a
divided cell containing a working electrode, a counter
electrode and a reference electrode generally produces
the best films. The commonly used anodes are
chromium, gold, nickel, palladium, titanium, platinum
and indium-tin oxide coated glass plates. Semi-conducting materials such as n-doped silicon (Noufi et al.,
1981), gallium arsenide (Noufi et al., 1980), cadmium
sulphide and semi-metal graphite (Bull et al., 1983) are
also used for the growth of polymer films. Electrochemical synthesis can be used to prepare free standing,
homogeneous and self doped films. Besides this, it is
possible to obtain copolymers and graft copolymers.
Polyazulene, polythiophene, polyaniline, polycarbazole
and several other polymers have been synthesized using
this approach.

1.5. Applications of conducting polymers

Fig. 1. Structures of some conducting polymers commonly used in

biosensors.

The increasing number of academic, governmental

and industrial laboratories throughout the world involved in basic research and assessment of possible
applications of conducting polymers show that this area
is interdisciplinary in nature. These conducting organic
molecular electronic materials have attracted much attention largely because of their many projected applications in solar cells, light weight batteries,

348

M. Gerard et al. / Biosensors & Bioelectronics 17 (2002) 345359

electrochromic devices, sensors and molecular electronic devices.

Polymeric heterojunctions, solar cells have been fabricated by electrochemical deposition of PPY on n-silicon (Audebert and Bidan, 1985). Many conducting
polymers such as polyacetylene, polythiophene, polyindole, polypyrrole, polyaniline etc. have been reported as
electrode materials for rechargeable batteries
(Saraswathi et al., 1999; Kawai et al., 1990; Santhanam
and Gupta, 1993). It has been reported (Gazard et al.,
1986) that the conducting polyheterocycles are good
candidates for electrochromic displays and thermal
smart windows. Scientists have used PPY films in a
neurotransmitter as a drug release system into the brain
(Zinger and Miller, 1984). The potential for conducting
polymers in the area of electronics and photonics (nonlinear optics) is enormous and has been used to fabricate diodes, capacitors, field-effect transistors (FET)
and printed circuit boards. Polyaniline PANI is being
used by Hitachi-Maxell for anti-static coating of 4 MB
barium ferrite floppy disk (Friend, 1993).
In analytical chemistry the problem of selectivity,
particularly at the low analyte concentrations and in
the presence of interfering substances is of paramount
importance. The development of sensors, which are
highly selective and easy to handle opens the door to
the problem in analysis. Conducting polymers have
enough scope for the development of various sensors. A
chemical or biosensor based on conducting polymers,
also rely on sensible changes in the optical and electrical properties of these materials. The industrial applications of biochemical and morphological processes in
fields such as production of pharmaceuticals, food
manufacturing, waste water treatment and energy production is on increase. This has led to the development
of biosensors. Biosensors have found promising applications in various fields such as biotechnology, food
and agriculture product processing, health care,
medicine and pollution monitoring.

2. Biosensors
The unprecedented interest in the development and
exploitation of analytical devices for detection, quantification and monitoring of specific chemical species has
led to the emergence of biosensors. The estimation of
metabolites such as glucose, urea, cholesterol and lactate in whole blood is of central importance in clinical
diagnostics. Biosensors represent a new trend emerging
in the diagnostic technology.
A biosensor is a device having a biological sensing
element either intimately connected to or integrated
within a transducer. The aim is to produce a digital
electronic signal, which is proportional to the concentration of a specific chemical or set of chemicals.

Fig. 2. Schematic of a simple biosensor.

Biosensor instruments are specific, rapid, simple to

operate, can be easily fabricated with minimal sample
pretreatment involved. The apparently alien marriage
of two contrasting disciplines combines the specificity
and sensitivity of biological systems with the computing
power of microprocessor. A general schematic of a
biosensor is shown in Fig. 2.
Depending on the level of integration, biosensors can
be divided into three generations. In the first generation, the biocatalyst is either bound to or entrapped in
a membrane, which in turn is fixed on the surface of the
transducer. The second generation biosensors involve
the adsorption of covalent fixation of the biologically
active component to the transducer surface and permits
the elimination of semi-permeable membrane. In the
case of third generation biosensors, it is the direct
binding of the biocatalyst to an electronic device that
transduces and amplifies the signal. Conducting polymer-based biosensors come in this category.

2.1. Transducer
The biochemical transducer or biocomponent imparts to the biosensor, selectivity or specificity. A transducer converts the biochemical signal to an electronic
signal. The transducer of an electrical device responds
in a way that a signal can be electronically amplified,
stored and displayed. Suitable transducing system can
be adapted in a sensor assembly depending on the
nature of the biochemical interaction with the species of
interest. The physical transducers vary from electrochemical, spectroscopic, thermal, piezoelectric and surface acoustic wave technology (Svorc et al., 1997;
Urban et al., 1990). The most common electrochemical
transducers being utilized are amperometric and
potentiometric.
An amperometric biosensor measures the current
produced during the oxidation or reduction of a
product or reactant usually at a constant applied potential. Such sensors have fast response times and good
sensitivity. However, the excellent specificity of the
biological component can be compromised by the partial selectivity of the electrode. This lack of specificity
requires sample preparation, separation or some compensation for interfacing signals. Potentiometric biosensors relate electrical potentials to the concentration of
analyte by using an ion-selective electrode or a gassensing electrode as the physical transducer (Lewenstam et al., 1994; Domansky et al., 1998). These are

M. Gerard et al. / Biosensors & Bioelectronics 17 (2002) 345359

selective, have large dynamic ranges and are nondestructive.

2.2. Biocomponents
Biocomponents, which function as biochemical transducers can be enzymes, tissues, bacteria, yeast, antibodies/antigens, liposomes, organelles (Sadik and Wallace,
1993; Foulds and Lowe, 1988; Schmid, 1987). Within a
biosensor the recognition biomolecule incorporated
possesses an exquisite level of selectivity but is vulnerable to extreme conditions such as temperature, pH and
ionic strength (Karyakin et al., 1999). Most of the
biological molecules such as enzymes, receptors, antibodies, cells etc. have very short lifetime in solution
phase. Thus they have to be fixed in a suitable matrix.
The immobilization of the biological component
against the environmental conditions results in decreased enzyme activity (Schuhmann et al., 1992; Evtugyn et al., 1998). The activity of immobilized
molecules depends upon surface area, porosity, hydrophillic character of immobilizing matrix, reaction
conditions and the methodology chosen for
immobilization.
A number of techniques such as physical adsorption,
cross-linking, gel entrapment, covalent coupling etc.
have been used to immobilize biological molecules in
carrier materials as shown in Table 1.
Various matrices have been used for the immobilization of enzymes such as membranes, gels, carbon,
graphite, silica, polymeric films etc. (Trojanowicz and
vel Krawczyk, 1995; Pandey, 1992; Ikeda et al., 1985;
Gun and Lev, 1996). There is thus a great need to
design the electrodes that are compatible with the biological component that can lead to rapid electron transfer at the electrode surface. Conducting polymers are
attractive as possible materials for such applications.

349

3. Importance of conducting polymers to biosensors

Conducting polymers have attracted much interest as
a suitable matrix of enzymes Conducting polymers are
used to enhance speed, sensitivity and versatility of
biosensors in diagnostics to measure vital analytes.
Conducting polymers are thus finding ever increasing
use in diagnostic medical reagents (Heller, 1990). In this
context several reviews on the use of conducting polymers in fabrication of efficient biosensors have been
published (Cosnier, 1999; Lewis et al., 1999; Kranz et
al., 1998; Santhanam, 1998; Trojanowicz et al., 1997;
Adeloju and Wallace, 1996; Trojanowicz and vel
Krawczyk, 1995; Alva and Phadke, 1994; Contractor et
al., 1994; Bartlett and Birkin, 1993; Schuhmann et al.,
1993; Deshpande and Amalnerkar, 1993; Boyle et al.,
1989; Bidan et al., 1988).
Conducting polymers have attracted much interest as
a suitable matrix for the entrapment of enzymes (Adeloju and Wallace, 1996; Bartlett and Cooper, 1993;
Sung and Bae, 2000). The techniques of incorporating
enzymes into electro-depositable conducting polymeric
films permit the localization of biologically active
molecules on electrodes of any size or geometry and is
particularly appropriate for the fabrication of multi-analyte micro-amperometric biosensors (Unwin and Bard,
1992). Electrically conducting polymers have considerable flexibility in the available chemical structure, which
can be modified as required. By chemical modeling and
synthesis, it is possible to modulate the required electronic and mechanical properties. Moreover, the polymer itself can be modified to bind protein molecules
(Lu et al., 1995; Mulchandani and Wang, 1996; Situmorang et al., 2000). Another advantage offered by
conducting polymers is that the electrochemical synthesis allows the direct deposition of the polymer on the

Table 1
Conventional immobilization procedures
Method

Advantages

Physical
adsorption

No modification of
biocatalyst. Matrix can be
regenerated. Low cost

Entrapment

Cross-linking

Disadvantages

Binding forces are

susceptible to change in pH,
temperature and ionic
strength
Only physical confinement of High diffusion barrier
biocatalyst near transducer.
Low cost

Loss of biocatalyst is
minimum. Moderate cost

Covalent bonding Low diffusional resistance

Stable under adverse
conditions

Harsh treatment of
biocatalyst by toxic
chemicals
Harsh treatment by toxic
chemicals. Matrix not
regenerable

Example

Reference

Adsorption of glucose
oxidase on conducting
polymers for glucose
detection
Entrapment of urease and
glutamate dehydrogenase in
polypyrrole/polyvinyl
sulphonate films for urea
detection
Glutaraldehyde mediated
linking of lactate
dehydrogenase for lactate
estimation
GOD binding via
poly(o-amino benzoic acid)
for glucose detection

Ramanathan 1995;
Ramanathan et al., 1996a,b

Gambhir et al., 2001b

Chaubey et al., 2000b

Ramanathan et al., 2000

350

M. Gerard et al. / Biosensors & Bioelectronics 17 (2002) 345359

electrode surface, while simultaneously trapping the

protein molecules (Bartlett and Whitaker, 1988;
Gambhir et al., 2001a). It is thus possible to control
the spatial distribution of the immobilized enzymes,
the film thickness and modulate the enzyme activity
by changing the state of the polymer. The development of any kind of technology in this field heavily
depends on the understanding of the interaction at
the molecular level, between the biologically active
protein, either as a simple composite or through
chemical grafting. For the proper relay of the electrons from the surface of the electrode to the enzyme
active site, the concept of electrical wiring has been
reported (Heller, 1990; Gregg and Heller, 1990). Conducting polymers are likely to provide a 3-dimensional electrically conducting structure for this
purpose.
Conducting polymers are also known to be compatible with biological molecules in neutral aqueous
solutions. They can be reversibly doped and undoped
electrochemically accompanied by significant changes
in conductivity and spectroscopic properties of the
films that can be used as a signal for the biochemical
reaction (Wrighton, 1986; Kittlesen et al., 1984). The
electronic conductivity of conducting polymers
changes over several orders of magnitude in response
to changes in pH and redox potential of their environment (Paul et al., 1985).
Conducting polymers have the ability to efficiently
transfer electric charge produced by the biochemical
reaction to electronic circuit (De Taxis du Poet et al.,
1990). Moreover conducting polymers can be deposited over defined areas of electrodes. This unique
property of conducting polymers along with the possibility to entrap enzymes during electrochemical
polymerization has been exploited for the fabrication
of amperometric biosensors (Umana and Waller,
1986; Foulds and Lowe, 1986; Iwakura et al., 1988;
Bartlett and Whitaker, 1988; Pandey, 1988). Besides
this, conducting polymers exhibit exchange and size
exclusion properties due to which they are highly sensitive and specific towards substrate (Price et al.,
1994; Mirmohseni et al., 1993; Teasdale and Wallace,
1993; Trojanowicz and vel Krawczyk, 1995). Numerous published papers reveal that the electrodeposition
of conducting polymers serve as good matrices for
the immobilization of enzymes (Trojanowicz and vel
Krawczyk, 1995; Kranz et al., 1998; Cooper and
Hall, 1992). They provide good detectability and fast
response as the redox reaction of the substrate, catalyzed by an appropriate enzyme, takes place in the
bulk of the polymer layer. Fig. 3 shows the pathway
suggested for electron transfer in the conducting polymer based amperometric biosensors.

Fig. 3. Pathway suggested for electron transfer in conducting polymer

based biosensors.

4. Immobilization of enzymes on conducting polymers

In recent years, numerous papers have been published indicating organic conducting polymers as a convenient
component,
forming
an
appropriate
environment for the immobilization of enzyme at the
electrode surface and its interaction with metallic or
carbon electrode surfaces. Stable immobilization of
macromolecular biomolecules on conducting microsurfaces with complete retention of their biological recognition properties is a crucial problem for the
commercial development of miniaturized biosensor.
Most of the conventional procedure for biomolecule
immobilization such as cross-linking, covalent binding
and entrapment in gels or membrane suffer from a low
reproducibility and a poor spatially controlled deposition. The other electrochemically prepared conducting
polymers used for the biomolecule immobilization are
polyacetylene, polythiophene, polyindole, polyaniline
etc. (Bartlett and Whitaker, 1987, 1988; Pandey, 1988;
Cooper and Hall, 1992). Also a few biosensors based
on insulating electropolymerized films polyphenol,
poly(o-phenylenediamine),
polydicholorophenolenedophenol and overoxidised polypyrrole have also been
elaborated (Malitesta et al., 1990; Bartlett and Caruana, 1992; Groom and Luong, 1993). Ramanathan
(1995a) have immobilized glucose oxidase (GOX) on
different conducting matrices such as polypyrrole
(PPY), polyaniline (PANI), polyaminobenzoic acid
(PAB) and have studied the response characteristics,
lifetime in detail. Ramanathan et al. (1996b) have also
studied the changes in the dielectric constant in PPY
immobilized GOD. Among the conducting polymers
PPY and its derivatives play a leading role due to their
versatile applicability and the wide variety of molecular
(redox) species covalently linked to a pyrrole group
(Bidan, 1992). Many recent reports have been published
on the immobilization of different enzymes into the
polyaniline film (Ramanathan et al., 1995b; Yang and
Shaolin, 1997; Chaubey et al., 2000a). Fig. 4 shows a
conducting polymer based screen printed electrode for
fabrication of amperometric biosensor.
A number of enzymes or biomolecules have been
immobilized physically on a number of conducting

M. Gerard et al. / Biosensors & Bioelectronics 17 (2002) 345359

polymers by (Ramanathan, 1995a; Ramanathan et al.,

1996b; Verghese et al., 1998; Gambhir et al., 2001b;
Chaubey et al., 2000a,b). This is the simplest method of
enzyme immobilization. The binding forces involved
are hydrogen bonds, multiple salt linkages, Vander
waals forces etc. Many enzymes may be incorporated
into conducting polymer films during electrochemical
deposition on appropriate electrodes. The entrapment
of enzymes in conducting polymers provides a facile
means for ensuring proximity between the active site of
the enzyme and the conducting surface of the electrode
with considerable potential for biosensor construction.
This method provides a facile and controllable method
for the deposition of biologically active molecules to
defined areas on the electrodes. For electrochemical
entrapment, a solution of the electro-polymerizable
monomer and the aqueous buffer containing the enzyme is used for the deposition process. The polymer
film can be deposited electrochemically using potentiostatic or galvanostatic mode. It has been established
that glucose oxidase can be successfully entrapped in
polypyrrole films (Umana and Waller, 1986; Foulds
and Lowe, 1986) and the morphology of the film alters
to a more fibrillar form. During electropolymerization
mediators can also be immobilized simultaneously with
enzyme (Iwakura et al., 1988; Sun and Tachikawa,
1992; Bartlett and Whitaker, 1987) but they can also be
conjugated with the monomer before polymerization.
This well controlled procedure of enzyme immobilization by electropolymerization is of great significance in
fabrication of microsensors in the preparation of multilayer devices (Sun and Tachikawa, 1992; Strike and de
Rooij, 1993) and multienzyme biosensor (Kajiya et al.,
1991; Yon-Hin et al., 1990; Yon-Hin and Lowe, 1992;
Tatsuma et al., 1993a,b; Garguilo et al., 1993). Many
theoretical models have also been coupled with the
electrochemical entrapment of enzyme for the evaluation of the role of the thickness of polymeric layers,
enzyme location, enzyme loading etc. on the biosensor
functioning (Bartlett and Whitaker, 1987; Gros and
Bergel, 1995; Gooding et al., 1998).

Fig. 4. A conducting polymer-screen printed electrode.

351

5. Types of biosensors

5.1. Amperometric biosensors

Amperometric biosensors measure the current produced during the oxidation or reduction of a product or
reactant usually at a constant applied potential. The
most important factor affecting the functioning of amperometric biosensors is the electron transfer between
catalytic molecule, usually oxidase or dehydrogenase,
and the electrode surface most often involving a mediation or conducting polymer. Although the role of the
electrodeposited conducting polymer films is not fully
understood and explained, various polymers used for
enzyme immobilization may affect significantly the response of biosensors sensitive to given species. Much
work has been done in recent years on the application
of electrochemically grown conducting polymer layers
in amperometric biosensors. Redox enzymes have been
either entrapped within conducting polymer layers or
covalently bound to functional groups (Bartlett and
Whitaker, 1987; Umana and Waller, 1986; Foulds and
Lowe, 1986; Fortier and Belanger, 1991; Hammerle et
al., 1992). Ramanathan et al. (1995c) have reported the
immobilization of GOD via manipulation of pore size
in PPY to improve loading parameters of enzyme leading to five fold increase in its current response. Verghese et al. (1998) have conducted similar experiments in
polyaniline/GOD films. Table 2 shows the different
biosensors developed for various substrates.

5.2. Potentiometric biosensors

Potentiometry is a rarely used detection method in
biosensor with enzymes immobilized in an electrodeposited polymer layer, although certain advantages
over amperometric detection for a PPY based electrode
with immobilized GOX have been demonstrated. For
biosensors having very slow response, the rate of potential change rather than steady state potential values,
should be considered as the analytical signal for quantification of the substrate (Trojanowicz and vel
Krawczyk, 1995).
Potentiometric biosensors with conducting polymers
can be produced using pH sensitivity of polymers (Ratcliffe, 1990). Polypyrrole sensitivity to NH3 was used to
produce such biosensors (Pandey and Mishra, 1988;
Trojanowicz et al., 1993). Conducting PPY molecular
interfaces have also been implemented to modulate
biological function of enzymes and living cells at the
electrode surface by adjustment of electrode potentials.
Often additionally obtained discrimination of electrochemical interfaces by using conducting polymers as
matrices for enzymes allows the use of such biosensors
for analysis of natural samples, e.g. flow injection determination of lactate in whole blood. Besides this, the

352

M. Gerard et al. / Biosensors & Bioelectronics 17 (2002) 345359

Table 2
Biosensors based on conducting polymers
Substrates or species to be
determined

Enzyme

Polymer

Detection

Glucose

Glucose oxidase

Polypyrrole

Amperometry
Potentiometry
Amperometry
Amperometry
Amperometry
Amperometry
Amperometry
Amperometry
Amperometry
Amperometry
Amperometry
Amperometry
Conductometry
Amperometry
Amperometry
Amperometry

Lipids
Phenols
Urea

Poly(N-methylpyrrole)
Polyaniline
Polyindole
Glucose dehydrogenase
Polypyrrole
Polypyrrole
D-amino acid oxidase
Tyrosinase
Polypyrrole
Cholesterol oxidase and Cholesterol esterase Polypyrrole
Choline oxidase
Substituted Polypyrrole
Glutamate dehydrogenase
Polypyrrole
Fructose dehydrogenase
Polypyrrole
Pepsin
Polyaniline
Lactate oxidase
Polyphenyline di-amine
Lactate dehydrogenase
Polyaniline
Polypyrrolepolyvinyl
sulphonate
Lipase
Polyaniline
Tyrosinase
Substituted polypyrrole
Urease
Polypyrrole

Uric acid
Triglycerides

Uricase
Lipase

D-Alanine
Atrazine
Cholesterol
Choline
Glutamate
Fructose
Hemoglobin
L-Lactate

advantage of well-defined formation of the polymer

layer, this methodology yields some difficulties, associated with a significant chemical and electrochemical
activity of the polymer matrix due to the similarity of
these materials towards ion-exchange processes and
redox equilibria. The interaction of NH3 with PPY was
utilized for a design of a potentiometric biosensor of
urea with urease immobilized in an electrodeposited
PPY layer. Potentiometric sensitivity was only 17 mV/
decade with strong interference in the presence of
potassium and sodium.
The creatinine electrode was made by co-immobilization of creatininase, creatinase and sarcosine oxidase in
a PPY matrix (Yamato et al., 1995). Difficulties concerning the immobilization of enzymes in the electrodeposited polymer layer or the mechanism of the
entrapment and dynamics effects on biosensors have
been widely discussed and till now no conclusive reports on the comparison of different polymer matrices
for immobilization of the same enzyme have been
related.

Polyaniline
Polyaniline

Conductometry
Amperometry
Amperometry
Potentiometry
Conductometry
Capacitance
Measurement
Admittance
measurement
Amperometry
Conductometry

5.3. Conductometric biosensors

Conductometric biosensors measure the changes in
the conductance of the biological component arising
between a pair of metal electrodes. Contractor et al.
(1994) have constructed biosensors for estimation of
glucose, urea neutral lipid/lipase and hemoglobin/
pepsin by monitoring the change in the electronic conductivity arising as a change in redox potential and/or
pH of the microenvironment in the polymer matrix.
Ramanathan et al. (1995b) have studied the application
of polyaniline LB films as a glucose biosensor. They
have also investigated the dielectric spectroscopic measurements of PPY/glucose biosensor (Ramanathan et
al., 1996b). Conductivity biosensors based on conducting polymers were developed for penicillin (Nishizawa
et al., 1992) and also for glucose, urea, lipids and
hemoglobin (Contractor et al., 1994). Microelectronic
devices have been fabricated with a sensitivity to urea
concentration in millimolar range in measurements of
conductance, capacitance and admittance using urease

M. Gerard et al. / Biosensors & Bioelectronics 17 (2002) 345359

immobilized in a PPY layer on gold film electrodes. A

flow injection system was developed with covalently
immobilized penicillinase on polypyrrole films and
changes in conductivity due to pH change was
measured.

5.4. Optical biosensors

Optical biosensors are based on the measurement of
light absorbed or emitted as consequence of a biochemical reaction. In such type of biosensors, light waves are
guided by means of optical fibers to suitable detectors.
These type of biosensors have been used for the detection of pH, O2 and CO2 etc. Gerard et al. (1999) have
measured the pyruvate concentration optically by immobilizing LDH onto polyaniline electrode. These electrodes were stable for 15 days and had a response time
of 90 s. Chaubey et al. (1998) have immobilized LDH
on PPY PVS electrodes on ITO plates and detected the
lactate concentration by optical measurements. Singhal
et al. (2000) immobilized glucose oxidase on polyhexyl
thiophene by LB technique and fabricated optical glucose sensor using UV visible spectroscopy.

5.5. Calorimetric biosensors

The basic principle of such biosensors is that all
biochemical reactions involve a change in enthalpy
Such a change in enthalpy is detected by calorimetric
biosensors. Mosbach and Danielsson (1981) developed
an enzyme thermister where the temperature change in
an enzyme reaction was measured by a thermister
device. A number of substrates, enzymes and antigens
have been estimated using thermister biosensors. Xie et
al. (1993) investigated a ferrocene mediated thermal
biosensor that combines electrochemistry and calorimetry. For detection, thermal signal generated by the
redox reaction was measured as opposed to measuring
the electrochemical signal.

5.6. Peizoelectric biosensors

These biosensors operate on the principle of generation of electric dipoles on subjecting an anisotropic
natural crystal to mechanical stress. Due to the adsorption of an analyte, the mass of the crystal is increased
resulting in altered frequency of oscillation. Such type
of biosensors have been utilized for the measurement of
ammonia, hydrogen, methane, carbon monoxide, nitrous oxide and other organophosphorous compounds.

6. Applications of conducting polymers

Conducting polymers have been used in the fabrication of biosensors in various fields such as:

353

Health care: In medical diagnosis (glucose, fructose,

lactate, ethanol, cholesterol, urea etc.)
Immunosensors: Can be used in medical diagnostics
and environmental sensors
DNA sensors: In the detection of various genetic
disorders.
Environmental monitoring: For control of pollution
and detection of hazardous chemicals in biosensors
(polyphenols, sulfites, peroxides, formaldehyde etc.)
Food analysis: For detection of glucose, fructose,
ethanol, sucrose, lactate, malate, galactose, citrate,
lactose, urea, starch etc. in food industries.

6.1. Biosensors for health care

Selective determination of various blood analytes viz.
glucose, urea, lactate, uric acid, cholesterol etc. is of
utmost importance for the screening and treatment of a
number of diseases.

6.1.1. Glucose biosensors

A number of reports on immobilization of glucose
oxidase in conducting polymers for glucose estimation
are available in literature (Fortier and Belanger, 1991;
Foulds and Lowe, 1986; Ramanathan, 1995a; Ramanathan et al., 1995b,c, 1996b; Umana and Waller,
1986). It has been investigated that the polymers containing para- and ortho-quinone groups as electrontransfer relay systems for oxido-reductases can
effectively catalyze the electro-oxidation of glucose. Ramanathan et al. (1995c) have immobilized GOD after
manipulation of the pore size in polypyrrole to improve
the loading of the enzyme. An exchange of para-toluene sulphonate and ferricyanide ions with a smaller ion
like chloride in solution has been applied for making
polypyrrole more porous enabling enhanced loading of
GOD. An attempt has been made to covalently couple
glucose oxidase (GOX) to poly(o-amino benzoic acid)
(PAB), a carboxy group functionalized polyaniline (Ramanathan et al., 2000). Mediated (with ferrocene carboxylic acid and tetrathiafulvalene) and unmediated
systems have been utilized for glucose concentrations.
6.1.2. Urea biosensors
Most of the urea biosensors available in literature are

based on detection of NH+

4 or HCO3 sensitive electrodes (Koncki et al., 2000; Hirose et al., 1983; Cho and
Huang, 1998; Pandey and Mishra, 1988; Gambhir et
al., 2001a). Osaka et al. (1999) constructed a highly
sensitive and rapid flow injection system for urea analysis with a composite film of electropolymerized inactive
polypyrrole and a polyion complex. Gambhir et al.
(2001a) have recently co-immobilized urease and glutamate dehydrogenase on electrochemically prepared
polypyrrole/polyvinyl sulphonate for the fabrication of
urea biosensor.

354

M. Gerard et al. / Biosensors & Bioelectronics 17 (2002) 345359

6.1.3. Lactate biosensors

Lactate biosensors are based on enzymes like lactate
oxidase and lactate dehydrogenase (Hart and Turner,
1996; Yang et al., 1999; Chaubey et al., 2000a,b Gerard
et al., 1999). Regeneration of lactate by substrate recycling has been utilized by co-immobilization of lactate
oxidase and lactate dehydrogenase by Chaubey et al.
(2000a). Recently lactate biosensor based on
polypyrrole/polyvinyl sulphonate composite films has
been reported by Chaubey et al. (2000b).
6.1.4. Cholesterol biosensors
Being a very important parameter in clinical diagnostics, cholesterol biosensors have attracted much attention. Amperometric cholesterol biosensors are based on
the enzyme cholesterol oxidase. Kajiya et al. (1991)
immobilized cholesterol oxidase and ferrocene carboxylate in polypyrrole electrochemically to describe the
sensitivity of the resulting films. Kumar et al. (2001) have
utilized dodecylbenzene sulphonate doped polypyrrole
films for immobilization of cholesterol oxidase by physical adsorption. These films were utilized for cholesterol
estimation via ferricyanide mediation.
6.2. Immunosensors
An effective combination of immunochemistry and
electrochemistry in an analytical device could provide
the basis of direct electrical detection of a wide range of
analytes with great sensitivity and specificity. In this
context, a number of immunosensors based on conducting polymers have been developed. A unique strategy for
reversible immunosensors using conducting polymers
based on pulsed electrochemical detection has been
developed by Sargents group. The strategy has been
utilized for the detection of organochlorine pesticides
including PCBs, atrazines and chlorinated phenols.
Porter (2000) investigated electroplated conducting polymers as antibody receptor in immunosensor. They have
shown that antibodies against conducting polymers (carbazole as a hapten) may react to modulate the polymer
electrochemistry. The reaction of the antiserum was
found to influence the polymer electrochemistry by an
amperometric response and therefore can be utilized as
a sensor for a direct immunoacid. AAI-Abtech product,
lead by Anthony Guiseppi-Elie is sensitive to H2O2 and
therefore may be used for enzyme, immunosensor, receptor and DNA probes and direct redox biosensor. The
company is marketing a polymer based system to monitor sulphate-reducing desulphovibrio bacteria, implicated in corrosion and biofouling. In this system, which
utilizes a disposable biosensor cartridge, antibodies specific to the bacterium are incorporated into an electroconductive polymer, which is deposited onto an array of
micro-electrodes on a silicon chip substrate. When the
electrodes are incubated with water samples, any

desulphovibrio bacteria present, may adhere to the

antibodies and result in a measurable change in conductivity. A company, Ohmicron Corporation, in Pennsylvania has also developed an immunosensor which is a
portable instrument for atrazine estimation. The instrument permits a total test time of less than 15 min and
can detect the pesticide at the level of parts per billion.

6.3. DNA Biosensors

DNA biosensors have an enormous application in
clinical diagnostics of inherited diseases, rapid detection
of pathogenic infections, and screening of cDNA
colonies required in molecular biology. Present methods
of genetic analysis require pre- and post-treatments to
modify DNAs with probes as proteins. Recently, DNA
integrated electroactive polymers (thin films or LB
monolayers) have provided intelligent materials which
possess superior intelligent material properties of self-assembly, self-multiplication, self-repair, self- degradation,
redundancy, and self-diagnosis (Minehan et al., 1994;
Garnier et al., 1999).
A few reports of interaction of DNA with conducting
polymers are available (Saoudi et al., 1997; Chehimi et
al., 1996; Bruno et al., 1994). Livache et al. (1995)
reported one-step electrodeposition of PPY films functionalized by a covalently linked oligonucleotide. Another possibility to dope DNA probes within
electropolymersised polypyrrole films and monitoring of
the current changes incurred by the hybridization (Wang
et al., 1999). Gambhir et al. (2001b) have reported the
results of the studies relating to the characteristics of
physically adsorbed DNA (Calf thymus) on conducting
PPY/PVS films. Immobilization of DNA on a conducting polymer matrix facilitates the detection of a signal
(amperometric or potentiometric) generated as a result
of interaction of proteins or drugs with DNA.

6.4. Biosensors for en6ironmental monitoring

Biosensors show a potential to complement both
laboratory based and field analytical methods for environmental monitoring (Marco and Barcelo, 1996;
Gabor, 2001; Lopez-Avila and Hill, 1997; Rogers, 1998).
Although a wide range of biosensors have been reported
for potential environmental applications, very few have
progressed into commercial markets. Biosensors with
potential for environmental applications have been the
subject of a number of recent reviews.
The conducting polymer based gas sensor for environmental monitoring is based on the fact that upon
exposure to vapor, the polymers show rapid conductivity
changes, which are generally reversible at room temperature. The change in conductivity probably results from
a reversible reduction of the polymer on exposure to the
organic vapor as well as change in the moisture content
of the film. (Harsanyi et al., 1999; Bartlett and LingChung, 1989a,b; Krutovertsev et al., 1992).

M. Gerard et al. / Biosensors & Bioelectronics 17 (2002) 345359

A thin film of conducting polymer based on microelectronic ammonia gas sensor device has been developed which is the first application of conducting
polymer in gas sensor devices for commercial applications. The device is a resistor element which shows large
variations when ammonia is present in the environment
in very low concentration and enables the necessary
action or alarm activation (www.ett.bme.hu).
Use of conducting polymers in impedance type sensors provide a possibility to build up low cost, highly
sensitive and selective room temperature gas sensor
(Bartlett et al., 1989c; Charlesworth et al., 1993). They
described a number of sensors for organic sensors for
organic vapours based on PPY, poly-N-methyl pyrrole,
poly(5-carboxy indole) and polyaniline. These materials
are however advantageous due to lack of specificity i.e.
they show responses to a wide range of different gases
and vapors. But their selectivity is found to be better
than that of the inorganic materials based gas sensitive
resistors.
AAI-Abtech, a small biotechnology company in
Pennsylvania, has developed a novel sensor based on
conducting PPY on an array of micoelectrodes. It has
been shown that a simple system using Mo(V)-catalyzed conversion of iodide to iodine can detect hydrogen peroxide. The polymer incorporates molybdenum
together with iodide, hydrogen peroxide converts the
iodide to iodine, which oxidizes the PPY, resulting in a
measurable change in conductivity.
Thomas Fare and coworkers have developed an immunosensor based on conducting polymer for screening
of pesticide atrazine. Adeloju and Yuan (1999) have
described a new approach for the flow amperometric
biosensing of formate with single, double and triple
layered arrangements of the ferrocyanide mediated
PPY based biosensor. The detection of formate concentration has been found to be important in atmosphere,
natural waters and sediments. This layered structure
comprised of formate dehydrogenase, NAD and mediator and has been demonstrated for the flow amperometric biosensing of formate in aqueous media, with the
detection limit of 2.5 mM.
A cost effective polyaniline base ammonia gas sensor
has been described by Lepsenyi et al. (1999). They
deposited sensing film of polyaniline, electrochemically
doped by cyclic voltammetry on thin and thick film
electrodes where the isolation gap is formed by laser
engraving. In this work, they investigated the sensitivity, selectivity, temperature, dependency and long term
behavior of the sensor.

6.5. Biosensors for food industry

Food industries need rapid and affordable methods
to determine compounds in the products for quality
control in food processing. In this context, the applica-

355

tion of biosensor technique may be an alternative to the

present inefficient and expensive methods. Extensive
work on development of analytical devices for estimation of glucose, sucrose, lactate, citric acid, ascorbic
acid etc. in the food products has been pursued during
the last few years (Pal et al., 1994; Stephens et al., 1998;
Stredansky et al., 1999). A few reports on the conducting polymers based biosensors for application to food
industries are available (Umana and Waller, 1986; Pal
et al., 1994; Chaubey et al., 2000a).
Ghosh (Hazra) et al. (1998) have developed a biosensor system for qualitative measurement of fish freshness. The biosensor is based on amperometric method
using three ferrocene carboxylic acid mediator incorporated conducting polypyrrole enzyme electrodes with
immobilized xanthine oxidase, nucleoside phosphorylase and nucleotidase enzymes for quantitative measurement of hypoxanthine, inosine and inosine
monophosphate metabolites in fish tissue.

7. Conclusion
Biochemical sensors have been shown to provide
complementary and additional information to that contributed by the well-established bioanalytical techniques. Particular advantages of biochemical sensors
concern the following: the possibility of miniaturizing
the setup, in principle down to the molecular scale, the
use of well-established microsystem technologies during
manufacture, integration of signal preprocessing steps
on a chip, and the building of arrays for more complex
pattern recognition analysis. By combining the use of
electronically conducting polymers with immobilized
enzymes and by making use of the particular properties
of conducting polymers, it is possible to develop novel
enzyme-based bioelectronic devices.

Acknowledgements
Authors are thankful to Dr. Krishon Lal, Director of
NPL for his interest in this field. Asha Chaubey is
grateful to CSIR, India for the award of senior research
fellowship.

References
Adeloju, S.B., Yuan, Y.J., 1999. Polypyrrole-based flow amperometric biosensor for formate, Electrochem. News, 4, online (http://
www.uws.edu.au).
Adeloju, S.B., Wallace, G.G, 1996. Conducting polymers and the
bioanalytical sciences. Analyst 121, 699 703.
Alva, S., Phadke, R.S., 1994. Conducting polymers in the fabrication
of efficient biosensors. Ind. J. Chem. 33A, 561 564.

356

M. Gerard et al. / Biosensors & Bioelectronics 17 (2002) 345359

Audebert, P., Bidan, G., 1985. Polyhalopyrroles: Electrochemical

synthesis and some characteristics. J. Electroanal. Chem. 190,
129 139.
Bartlett, P.N., Birkin, P.R., 1993. The application of conducting
polymers in biosensors. Synth. Met. 61, 15 21.
Bartlett, P.N., Whitaker, R.G., 1988. Strategies for the development
of amperometric enzyme electrodes. Biosensors 3, 359 379.
Bartlett, P.N., Cooper, J.M., 1993. A review of the immobilization of
enzymes in electropolymerized films. J. Electroanal. Chem. 362,
112.
Bartlett, P.N., Ling-Chung, S.K., 1989a. Conducting polymer gas
sensors part II: response of polypyrrole to methanol vapour. Sens.
Actat. B 19, 141 150.
Bartlett, P.N., Ling-Chung, S.K., 1989b. Conducting polymer gas
sensors Part II: results for 4 different polymers and 5 different
vapours. Sens. Actuat B. 20, 287 292.
Bartlett, P.N., Archer, PBM, Ling-Chung, S.K., 1989c. Conducting
polymer gas sensors part I: fabrication and characterization. Sens.
Actuat. 19, 125 140.
Bartlett, P.N., Caruana, D.J., 1992. Electrochemical immobilizaion of
enzymes. Part V. Microelectrodes for the detection of glucose
based on glucose oxidase immobilized in a poly(phenol) film.
Analyst 117, 1287 1292.
Bartlett, P.N., Whitaker, R.G., 1987. Electrochemical immobilization
of enzymes. Part II. Glucose oxidase immobilized in poly-Nmethyl pyrrole. J. Electroanal. Chem. 224, 37 48.
Bidan, G., 1992. Electroconducting conjugated polymers. New sensitive matrices to build up chemical or electrochemical sensors. A
Review. Sens. Actuat. B 6, 45 56.
Bidan, G., Ehui, B., Lapkowski, M., 1988. Conductive polymers with
immobilized dopants: ionomer composites and auto-doped polymers a review and recent advances. J. Apys. D. Appl. Phys. 21,
1043 1054.
Bloor, D., Movaghar, B., 1983. Conducting polymers. IEEE Proceedings 130, 225 232.
Boyle, A., Genies, E.M., Lapkowski, M., 1989. Application of electronic conducting polymers as sensors: polyaniline in the solid
state for detection of solvent vapours and polypyrrole for detection of biological ions in solutions. Synth. Metals 28, C769 C774.
Bruno, F.F., Marx, K.A., Tripathi, S.K., Akkara, J.A., Samuelson,
L.A., Kaplan, D.L., 1994. Enzyme mediated polymerization of
phenol and aniline derivatives on a langmuir trough to form
ordered 2-D polymer films. J. Int. Mat. Sys. Struct. 5, 631 634.
Bull, R.A., Fan, F.R., Bard, A.J., 1983. Polymer films on electrodes.
J. Electrochem. Soc. 130, 1636 1638.
Charlesworth, J.M., Partridge, A.C., Garrard, N., 1993. Mechanistic
studies in the interactions between polypyrrole and organic vapours. J. Phys. Chem 97, 5418 5423.
Chaubey, A., Pande, K.K., Singh, V.S., Malhotra, B.D., 2000a.
Co-immobilization of lactate oxidase and lactate dehydrogenase
on conducting polyaniline films. Anal. Chim. Acta 407, 97 103.
Chaubey, A., Gerard, M., Singhal, R., Singh, V.S., Malhotra, B.D.,
2000b. Immobilization of lactate dehydrogenase on electrochemically prepared polypyrrole-polyvinyl sulphonate composite films
for application to lactate biosensors. Electrochim. Acta 46, 723
729.
Chaubey, A., Singhal, R., Gerard, M., Malhotra, B.D., 1998. Lactate
biosensor based on polypyrrole-polyvinylsulphonate composites.
Proceedings of the 6th Asian Conference on Solid State Ionics:
Science and Technology, World Scientific Publishing Co. Ltd,
Singapore, pp. 479 483.
Chehimi, M.M., Abel, M.L., Saoudi, B., Delamar, M., Jammul, N.,
Watts, J.F., Zhdan, P.A., 1996. Adsorption of macromolecules
onto conducting polymers. Polymery 41, 75 84.
Cho, W.J., Huang, H.J., 1998. An amperometric urea biosensor
based on a polyaniline-perfluorosulphonated ionomer composite
electrode. Anal. Chem. 70, 3946 3951.

Contractor, A.Q., Sureshkumar, T.N., Narayanan, R, Sukeerthi, S.,

Lal, R., Srinivasa, R.S., 1994. Conducting polymars based biosensors. Electrochim. Acta 39, 1321 1324.
Cooper, J.C., Hall, E.A.H., 1992. Electrochemical response of an
enzyme-loaded polyaniline film. Biosens. Bioelectron. 7, 473 485.
Cosnier, S., 1999. Biomolecule immobilization on electrode surfaces
by entrapment or attachment to electrochemically polymerized
films. A review. Biosens. Bioelectron 14, 443 456.
De Taxis du Poet, P., Miyamoto, S., Murakami, T., Kimura, J.,
Karube, I., 1990. Direct electron transfer with glucose oxidase
immobilized in an electropolymerized poly-N-methylpyrrole film
on a gold microelectrode. Anal. Chim. Acta 235, 255 264.
Deshpande, M.V., Amalnerkar, D.P., 1993. Biosensors prepared from
electrochemically synthesized conducting polymers. Prog. Polym.
Sci 18, 623 649.
Domansky, K., Baldwin, D.L., Grate, J.W., Hall, T.B., Li, J., Josowicz, M., Janata, J., 1998. Development and calibration of field
effect transistor-based sensor array for measurement of hydrogen
and ammonia gas mixtures in humid air. Anal. Chem. 70, 473
481.
Duke, C.B., Schein, L.B., 1980. Organic solids: is energy-based theory
enough? Phys. Today 33, 42 48.
Evtugyn, G.A, Budnikov, H.C., Nikolskaya, E.B., 1998. Sensitivity
and selectivity of electrochemical enzyme sensors for inhibitor
determination. Talanta 46, 465 484.
Fortier, G., Belanger, D., 1991. Characterization of the biochemical
behavior of glucose oxidase entrapped in a polypyrrole film.
Biotechnol. Bioengg. 37, 854 858.
Foulds, N.C., Lowe, C.R., 1986. Enzyme entrapment in electrically
conducting polymers. J. Chem. Soc. Faraday Trans. I 82, 1259
1264.
Foulds, N.C., Lowe, C.R., 1988. Immobilization of glucose oxidase in
ferrocene modified pyrrole polymers. Anal. Chem 60, 2473 2478.
Friend, R.H. (Ed.) 1993. Conductive Polymer II, Rapra Review
Report, 6 (3), 23.
Gabor, H., (2001). Highly sensitive and selective ammonia sensor for
monitoring and workplace safety applications, (www.ett.bme.hu).
Gambhir, A. Gerard, M., Jain, S.K., Malhotra, B.D., 2001a. Characterization of DNA immobilized on electrochemically prepared
conducting polypyrrole-polyvinyl sulphonate films. Appl.
Biochem. Biotech. (in press).
Gambhir, A., Gerard, M., Mulchandani, A., Malhotra, B.D., 2001b.
Co-immobilization of urease and glutamate dehydrogenase in
electrochemically prepared polypyrrole-polyvinyl sulphonate
films, Appl. Biochem. Biotechnol. (in press).
Garguilo, M.G., Huynh, N., Proctor, A., Michael, A.C., 1993. Amperometric sensors for peroxide, choline and acetylcholine based
on electron transfer between horseradish peroxidase and a redox
polymer. Anal. Chem. 65, 523 528.
Garnier, F., Korri-Youssoufi, H., Srivastava, P., Mandrand, B.,
Delair, T., 1999. Toward intelligent polymers: DNA sensors based
on oligonucleotide-functionalized polypyrroles. Synthetic Metals
100, 89 94.
Gazard, M., 1986. In: Skotheim, T.A. (Ed.), Handbook of Conducting Polymers, vol. 1. Marcel Dekker, USA, p. 673.
Gerard, M., Ramanathan, K., Chaubey, A., Malhotra, B.D., 1999.
Immobilization of lactate dehydrogenase on electrochemically
prepared polyaniline films. Electroanalysis 12, 450 452.
Ghosh (Hazra), S., Sarker, D., Misra, T.N., 1998. Development of an
amperometric enzyme electrode biosensor for fish freshness detection. Sens. and Actat. B 53, 58 62.
Gooding, J.J., Hall, E.A.H, Hibbert, D.B., 1998. Thick film to
monolayer recognition layers in enzyme elcetrodes. Electroanalysis 10, 1130 1136.
Greene, R.L., Street, G.B., Suter, L.J., 1975. Superconductivity in
polysulfur nitride (SN)x. Phys. Rev. Lett. 34, 577 579.

M. Gerard et al. / Biosensors & Bioelectronics 17 (2002) 345359

Gregg, B.A., Heller, A., 1990. Cross-linked redox gels containing
glucose oxidase for amperometric biosensor applications. Anal.
Chem. 62, 258 263.
Groom, C.A., Luong, J.H.T., 1993. Improvement of the selectivity of
amperometric biosensors by using a permselective electropolymerized film. Anal. Lett. 26, 1383.
Gros, P., Bergel, A., 1995. Improved model of a polypyrrole glucose
oxidase modified electrode. J. Electroanal. Chem. 386, 65 73.
Guiseppi-Elie, A., Wallace, G.G., Matsue, T., 1997. Chemical and
biological sensors based on electrically conducting polymers. In:
Skotheim, T., Elsenbaumer, R., Reynolds, J.R. (Eds.), Handbook
of Conducting Polymers, 2nd ed. Marcel Dekker, New York, pp.
963 991 Chapter 34.
Gun, J., Lev, O, 1996. Sol-gel derived ferrocenyl-modified silicategraphite composite electrode: wiring of glucose oxidase. Anal.
Chim. Acta 336, 95 106.
Hammerle, M., Schuhmann, W., Schmidt, H.L., 1992. Amperometric
polypyrrole enzyme electrodes: effect of permeability and enzyme
location. Sens. Actuat. B 6, 106 112.
Harsanyi, G., Dobay, R., Visy, Cs., 1999. Conducting polymer based
electrochemical sensors on thick film substrate. Electroanalysis 11,
804 808.
Hart, A.L., Turner, A.P.F., 1996. On the use of screen and ink-jet
printing to produce amperometric enzyme electrodes for lactate.
Biosens. Bioelectron 11, 263 270.
Heeger, A.J., 1986. In: Skotheim, T.A. (Ed.), Handbook of Conducting Polymers, vol. II. Marcel Dekker, New York, p. 729 and
references therein.
Heller, A., 1990. Electrical wiring of redox and enzymes. Acc. Chem.
Res. 23, 128 134.
Hirose, S., Hayashi, M, Tamura, N., Kamidate 1983. Determination
of urea in blood serum with use of immobilized urease and a
microwave cavity ammonia monitor, Anal. Chim. Acta, 151,
377 382.
Ikeda, T., Hamad, H., Miki, K.and, Senda, M., 1985. Glucose
oxidase immobilized benzoquinone-carbon paste electrode as a
glucose sensor. Agric. Biol. Chem. 49, 541 543.
Ivory, D.M., Miller, G.G., Sowa, J.M., Shacklette, L.W., Chance,
R.R., and. Baughman, R.H, 1979. Highly conducting chargetransfer complexes of poly(p-phenylene). J. Chem. Phys. 71,
1506 1507.
Iwakura, C., Kajiya, Y., Yoneyama, H., 1988. Simultaneous immobilization of glucose oxidase and a mediator in conducting polymer
films. J. Chem. Soc. Chem. Commun., 1019 1020.
Kajiya, Y., Sugai, H., Iwakura, C., Yoneyama, H., 1991. Glucose
sensitivity of polypyrrole films containing immobilized glucose
oxidase and hydroquinone sulphonate ions. Anal. Chem. 63,
49 54.
Kanazawa, K.K., Diaz, A.F., Geiss, R.H., Gill, W.D., Kwak, J.F.,
Logan, J.A., Rabolt, J.F., Street, G.B., 1979. Organic Metals:
polypyrrole, a stable synthetic metallic polymer. J. Chem. Soc.
Chem. Commun., 854855.
Karyakin, A.A., Vuki, M., Lukachova, L.V., Karyakina, E.E., Orlov,
A.V., Karpachova, G.P., Wang, J., 1999. Processable polyaniline
as an advanced potentiometric pH transducers. Anal. Chem. 71,
2534 2540.
Kawai, T., Kuwabara, T., Wang, S., Yoshino, K., 1990. Secondary
battery characteristics of poly(3-alkylthiophene). Jap. J. Appl.
Phys. 29, 602 605.
Kittlesen, G.P., White, H.S., Wrighton, M.S., 1984. Chemical derivatization of microelectrode arrays by oxidation of pyrrole and
N-methyl pyrrole. Fabrication of molecule-based electronic
devices. J. Am. Chem. Soc. 106, 7389 7396.
Koncki, R., Radomska, A., Glab, S., 2000. Potentiometric determination of dialysate urea nitrogen. Talanta 52, 13 17.
Kranz, C., Wohlschlager, H., Schmidt, H.L., Schuhmann, W., 1998.
Controlled electrochemical preparation of amperometric biosen-

357

sor based on conducting polymer multilayers. Electroanalysis 10,

546 552.
Kroschwitz, J.I., 1988. In: Kroschwitz, J.I. (Ed.), Electrical and
Electronic Properties of Polymers. Wiley, New York.
Krutovertsev, S.A., Sorokin, S.I., Zorin, A.V., Letuchy, Y.A., Antonova, O.Y., 1992. Polymer film based sensors for ammonia
detection. Sens. and Actuat. B 7, 492 494.
Kumar, A., Rajesh, Chaubey, A., Grover, S.K., Malhotra, B.D.,
2001. Immobilization of cholesterol oxidase and potassium ferricyanide on dodecylbenzene sulfonate ion doped polypyrrole film.
J. Appl. Polym. Sci. 82, 3486 3491
Lepsenyi, I, Reichardt, A., Inzelt, G., Harsanyi, G., 1999. Highly
sensitive and selective polymer based gas sensors, Proc. of twelfth
European microelectronics and packaging Conf., Harrogate, UK,
pp. 301 305.
Lewenstam, A., Babacka, J., Ivaska, A., 1994. Mechanism of ionic
and redox sensitivity of p-type conducting polymers. Part I.
Theory. J. Electroanl. Chem. 368, 23 31.
Lewis, T.W., Wallace, G.G., Smyth, M.R., 1999. Electrofunctional
polymers: their role in the development of new analytical systems.
Analyst 124, 213 219.
Livache, T., Roget, A., Dejean, E., Barthet, C., Bidan, G., Teoule,
R., 1995. Biosensing effects in functionalized electroconducting
conjugated polymer layers: addressable DNA matrix for the detection of gene mutations. Synth. Met. 71, 2143 2146.
Lopez-Avila, V., Hill, H.H., 1997. Field analytical chemistry. Anal.
Chem. 69, 289R 305R.
Lu, W., Zhao, H., Wallace, G.G., 1995. Pulsed electrochemical
detection of proteins using conducting polymer sensors. Anal.
Chim. Acta 315, 27 32.
Malitesta, C., Palmisano, F., Torsi, L., Zambonin, P.G., 1990. Glucose fast-response amperometric sensor based on glucose oxidase
immobilized in an electropolymerized poly(o-phenylenediamine)
film. Anal. Chem. 62, 2735 2740.
Marco, M.P., Barcelo, D., 1996. Environment applications of analytical biosensors. Measurement Sci Technol. 7, 1547 1572.
Minehan, S., Marx, K.A., Tripathi, S.K., 1994. Kinetics of DNA
binding to electrically conducting polypyrrole films. Macromolecules 27, 777 783.
Minot, M.J., Peristein, J.H., 1971. Mixed valence square planar
complexes: a new class of solids with high electrical conductivity
in one dimension. Phys. Rev. Lett. 26, 371 373.
Mirmohseni, A., Price, W.E., Wallace, G.G., 1993. Electrochemically
controlled transport across conducting polymer composites basis of smart membrane materials. Polymer Gels and Networks 1,
61 77.
Mosbach, K, Danielsson, B, 1981. Thermal bioanalyzers in flow
streams enzyme thermister devices. Anal. Chem. 53, 83A 94A.
Mulchandani, A., Wang, C.L., 1996. Bienzyme sensors based on
poly(anilinomethylferrocene)-modified electrode. Electroanalysis
8, 414 419.
Nishizawa, M., Matsue, T., Uchida, I., 1992. Penicillin sensor based
on a microarray electrode coated with pH-responsive polypyrrole.
Anal. Chem. 64, 2642 2644.
Noufi, R., Frank, A.J., Nozik, A.J, 1981. Stabilization of n-type
silicon photoelectrodes to surface oxidation in aqueous electrolyte
solution and mediation of oxidation reaction by surface attached
organic conducting polymers. J. Am. Chem. Soc. 103, 1849 1850.
Noufi, R., Tench, D., Warren, L.F., 1980. Protection of n-GaAs
photoanodes with photoelectrochemically generated polypyrrole
films. J. Electrochem. Soc. 127, 2310 2311.
Osaka, T., Komaba, S., Fujino, Y., Matsuda, T., Satoh, I., 1999.
High sensitivity flow injection analysis of urea using composite
electropolymerized polypyrrole-polyion complex film. J. Electrochem Soc. 146, 615 619.
Pal, P., Nandy, D., Misra, T.N., 1994. Immobilization of alcohol
dehydrogenase enzyme in a Langmuir Blodgett film of stearic

358

M. Gerard et al. / Biosensors & Bioelectronics 17 (2002) 345359

acid. Its application as an ethanol sensor. Thin Solid Films 234,

138 143.
Pandey, P.C., 1992. Membrane electrodes as biosensor. Bull. Electrochem. 8, 212 221.
Pandey, P.C., 1988. A new conducting polymer-coated glucose sensor. J. Chem. Soc. Faraday Trans. I 84, 2259 2265.
Pandey, P.C., Mishra, A.P., 1988. Conducting polymer-coated enzyme microsensor for urea. Analyst 113, 329 331.
Paul, E.W., Ricco, A.J., Wrighton, M.S., 1985. Resistance of polyaniline films as a function of electrochemical potential and the
fabrication of polyaniline-based microelectronic devices. J. Phys.
Chem. 89, 1441 1447.
Porter, R.A., 2000. Investigation of electroplated conducting polymers as antibody receptors in immunosensors. J. Immunoassay
21, 51 64.
Price, W.E., Wallace, G.G., Zhao, H., 1994. Effect of counter ion on
transport across polypyrrole membranes. J. Membr. Sci. 87, 47
56.
Rabolt, J.F., Clarke, T.C., Kanazawa, K.K., Reynolds, J.R., Street,
G.B., 1980. Organic metals: poly(p-phenylene sulphide) hexafluoroarsenate. J. Chem. Soc. Chem. Commun., 347 348.
Ramanathan, K., 1995a. Application of some conducting polymers to
biosensor, Ph.D Thesis, IIT Delhi, India.
Ramanathan, K., Ram, M.K., Malhotra, B.D., Murthy, A.S.N,
1995b. Application of polyaniline-Langmuir-Blodgett films as a
glucose biosensor. Mater. Sci. Engg. C 3, 159 163.
Ramanathan, K., Mehrotra, R., Jayaram, B., Murthy, A.S.N., Malhotra, B.D, 1996a. Simulation of electrochemical process for
glucose oxidase immobilized conducting polymer. Anal. Lett. 29,
1477 1484.
Ramanathan, K., Ram, M.K., Verghese, M.M., Malhotra, B.D.,
1996b. Dielectric spectroscopic studies on polypyrrole glucose
oxidase films. J. Appl. Polym. Sci. 60, 2309 2316.
Ramanathan, K., Sundaresan, N.S., Malhotra, B.D., 1995c. Ion
exchanged polypyrrole-based glucose biosensor: enhanced loading
and response. Electroanalysis 7, 579 582.
Ramanathan, K., Pandey, S.S., Kumar, R., Gulati, A., Murthy,
A.S.N., Malhotra, B.D., 2000. Covalent immobilization of glucose oxidase to poly(o-amino benzoic acid) for application to
glucose biosensor. J. Appl. Polym. Sci. 78, 662 667.
Ratcliffe, N.M., 1990. Polypyrrole-based sensor for hydrazine and
ammonia. Anal. Chim. Acta 239, 257 262.
Rogers, K.R., 1998. Biosensor technology for environment measurement. In: Meyers, R.A. (Ed.), Encyclopedia of Environmental
Analysis and Remediation. John Wiley and Sons, New York, pp.
755 768.
Sadik, O., Wallace, G.G, 1993. Pulsed amperometric detection of
proteins using antibody containing conducting polymers. Anal.
Chim. Acta 279, 209 212.
Santhanam, K.S.V., 1998. Conducting polymers for biosensors: rationale based on models. Pure Appl. Chem 70, 1259 1262.
Santhanam, K.S.V., Gupta, N., 1993. Conducting-polymer electrodes
in batteries. TRIP 1, 284 289.
Saoudi, B., Jammul, N., Abel, M.L., Chehimi, M.M., Dodin, G.,
1997. DNA adsorption onto conducting polypyrrole. Synth. Met.
87, 97 103.
Saraswathi, R., Gerard, M., Malhotra, B.D., 1999. Characteristics of
aqueous polycarbazole batteries. J. Appl. Polym. Sci. 74, 145
150.
Schmid, R.D., 1987. Preface, in Biosensors International Workshop,
R.D. Schmid, (Ed.), 10, VCH, Weinheim, Federal Republic of
Germany.
Schuhmann, W., 1995. Conducting polymers and their application in
amperometric biosensors. Mikrochim. Acta 121, 1 29.
Schuhmann, W., Kranz, C., Huber, J., Wohlschlager, H., 1993.
Conducting polymer-based amperometric enzyme electrodes. Towards the development of miniaturized reagentless biosensors.
Synth. Metals 61, 31 35.

Schuhmann, W., Lehn, C, Schmidt, H.L, Grundig, B., 1992. Comparison of native and chemically stabilized enzymes in amperometric enzyme electrodes. Sens. and Actuat. B 7, 393 398.
Shirakawa, H., Louis, E.J., MacDiarmid, A.G., Chiang, C.K.,
Heeger, A.J., 1977. Synthesis of electrically conducting organic
polymers: halogen derivatives of polyacetylene, (CH)x. J. Chem.
Soc. Chem .Commun., 578.
Singhal, R., Chaubey, A., Pande, K.K., Malhotra, B.D., 2000. Immobilization of Glucose oxidase on Poly(3-Hexyl Thiophene) LB
films for application to Glucose biosensor in 4th National Conference on Solid State Ionics, 3 5 March, 2000 at Indian Institute of
Technology, Mumbai.
Situmorang, M., Hibbert, D.B., Gooding, J.J., 2000. An experimental
design study of interferences of clinical relevance of polytyramine
immobilized enzyme biosensors. Electroanalysis 12, 111 119.
Situmorang, M., Gooding, J.J., Hibbert, D.B., Barnett, D., 1998.
Electrodeposited polytyramine as an immobilization matrix for
enzyme biosensors. Biosens. Bioelectron. 13, 953 962.
Stephens, S.K., Cullen, D.C., Warner, P.J., 1998. Biosensors for
novel rapid assay methods in the food industry. New Food 1,
47 51.
Stredansky, M, Pizzariello, A., Stredanska, S., Miertus, S., 1999.
Determination of D-fructose in food stuffs by an improved amperometric biosensor based on a solid binding matrix. Anal. Commun. 36, 57 61.
Strike, D.J., de Rooij, N.F., Koudelka-Hep, M., 1993. Electrodeposition of glucose oxidase for the fabrication of miniature sensors.
Sens. Actuat. B 13, 61 64.
Sun, Z., Tachikawa, H., 1992. Enzyme-based bilayer conducting
polymer electrodes consisting of polymetallopthalocyanines and
polypyrrole-glucose oxidase thin films. Anal. Chem. 64, 1112
1117.
Sung, W.J., Bae, Y.H., 2000. A glucose oxidase electrode based on
electropolymerized conducting polymer with polyanion-enzyme
conjugated dopant. Anal. Chem. 72, 2177 2181.
Svorc, J., Miertu, S., Katrlik, J., Stredansk, M., 1997. Composite
transducer for amperometric biosensors. The glucose sensor.
Anal. Chem 69, 2086 2089.
Tatsuma, T., Watanabe, T., Watanabe, T., 1993a. Polypyrrole bioenzyme electrodes with glucose oxidase and peroxidase. Sens. Actuat. B 14, 752 753.
Tatsuma, T., Watanabe, T., Watanabe, T., 1993b. Electrochemical
characterization of polypyrrole bioenzyme electrodes with glucose
oxidase and peroxidase. J. Electroanal. Chem. 356, 245 253.
Teasdale, P.R., Wallace, G.G., 1993. Molecular recognition on conducting polymers: basis of electrochemical sensing technology.
Analyst 118, 329 334.
Tourillon, G., Garnier, F., 1983. Stability of conducting polythiophene and derivatives. J. Electrochem. Soc. 130, 2042 2044.
Trojanowicz, M., vel Krawczyk, T.K., 1995. Electrochemical biosensors based on enzymes immobilized in electropolymerized films.
Mikrochim. Acta 121, 167 181.
Trojanowicz, M., vel Krawczyk, T.K., Alexander, P.W., 1997. Organic conductings as active materials in electrochemical chemosensors and biosensors. Chem. Anal. 42, 199 213.
Trojanowicz, M., Matuszewski, W., Szczepanczyk, B., Lewenstam,
A., 1993. In: Guilbault, G.G., Mascini, M. (Eds.), Uses of Immobilized Biological Compounds. Kluwer, Dordrecht, pp. 141 150.
Umana, M., Waller, J., 1986. Protein modified electrodes: the glucose
oxidase/polypyrrole system. Anal. Chem. 58, 2979 2983.
Unwin, P.R., Bard, A.J., 1992. Ultramicroelectrode voltammetry in a
drop of solution: a new approach to the measurement of adsorption isotherms at solid liquid interface. Anal. Chem. 64, 113
119.
Urban, G., Jachimowicz, A., Kohl, F., Kuttner, H., Olcaytug, F.,
Kamper, H., Pittner, F., Mann-Buxbaum, E., Schalkhammer, T.,
Prohaska, O., Schonauer, M., 1990. High resolution thin film

M. Gerard et al. / Biosensors & Bioelectronics 17 (2002) 345359

temperature sensor arrays for medical applications. Sens. Actuat.
A 21 23, 650 654.
Verghese, M.M., Ramanathan, K., Ashraf, S.M., Malhotra, B.D.,
1998. Enhanced loading of glucose oxidase on polyaniline films
based on anion exchange. J. Appl. Polm. Sci. 70, 1447 1453.
Wang, J., Jiang, M., Aantonio, F., Mukherjee, B, 1999. New labelfree DNA recognition based on doping nucleic acid probes within
conducting polymer films. Anal. Chim. Acta 402, 7 12.
Wrighton, M.S., 1986. Surface functionalization of electrodes with
molecular reagents. Science 231, 32 37.
Wring, S.A, Hart, J.P., 1992. Chemically modified carbon-based
electrodes and their application as electrochemical sensors for the
analysis of biologically important compounds. Analyst 117,
1215 1229.
Xie, B., Khayyami, M., Nwosu, T., Larsson, P., Danielsson, B., 1993.
Ferrocene mediated thermal biosensor. Analyst 118, 845 848.

359

Yamato, H., Ohawa, M.and, Wernet, W., 1995. A polypyrrole/three

enzyme electrode for creatinine detection. Anal. Chem. 67, 2776
2780.
Yang, Q., Atanasov, P., Wilkins, E., 1999. Needle type lactate
biosensor. Biosens. Bioelectron. 14, 203 210.
Yang, Y., Shaolin, M., 1997. Bioelectrochemical responses of
polyaniline horseradish peroxidase electrodes. J. Electroanal.
Chem. 432, 71 78.
Yon Hin, B.F.Y., Lowe, C.R., 1992. Amperometric response of
polypyrrole entrapped bienzyme films. Sens. Actuat. B 7, 339
342.
Yon-Hin, B.F.Y., Sethi, R.S., Lowe, C.R., 1990. Multianalyte microelectronic biosensors. Sens. Actuat. B 1, 550 554.
Zinger, B., Miller, L.L., 1984. Timed release of chemicals from
polypyrrole films. J. Am. Chem. Soc. 106, 6861 6863.