6th Pan-Hellenic Conference on Biomedical Technology (ELEVIT 2015)

ELECTROCHEMICAL BIOSENSORS BASED ON PROTEIN

IMMOBILIZATION ON MESOPOROUS TiO2 FILMS
C. Tiflidis, S. Katsiaounis, C. Tsekoura and E. Topoglidis*

* Department of Materials Science, University of Patras, Rio 26504, Greece

* etop@upatras.gr

Introduction ranges of H2O2 on the different prepared TiO 2 films are

The immobilization of biomolecules upon electrodes presented in table 1.
and other surfaces is important not only for studying
their function and structure but also for the development Table 1: Sample codes and H2O2 biosensing sensitivity
of bioanalytical devices such as biosensors. We of different prepared TiO2 films.
investigate in this study the use of various thin Scan Rate Detection
Electrode vs. Ag/AgCl
mesoporous TiO2 films as substrates for protein (mV/s) Range (M)
immobilization.
TiO2 SG/Cyt-c 25 20-320
Mesoporous TiO2 films have a high surface area, are TiO2 P25/Cyt-c 25 140-530
optically transparent, exhibit good electrical
conductivity and can be prepared by low cost screen TiO2 P25 (triton-X)/Cyt-c 50 100-400
printing technologies. Therefore they provide a novel
and versatile immobilization surface for the
development of optical and electrochemical biosensors
based on the reactivity of the immobilized proteins. [1]

Materials and Methods

For this study, a small redox protein, Cytochrome-c
(Cyt-c), has been successfully immobilized on the
surface of mesoporous TiO2 films prepared by the Sol-
Gel or the suspension methods (TiO 2-SG, TiO2-P25 and
TiO2-P25 mixed with Triton-X and HNO 3, all films
annealed at 450oC).
Results and Discussion
This approach is particularly attractive, because not only
it allows high protein loading in a stable and functional
way but also allows the function of the immobilized
protein to be investigated optically, electrochemically
and spectroelectrochemically without the addition of
any electron-transfer mediators or promoters. [2]
Therefore in this study we have examined the potential
of these TiO2 films to be used for the development of an
electrochemical H2O2 biosensor by initially
characterizing their surface using a variety of techniques
such as SEM, TEM and XRD.
Furthermore we have examined the optical and
electrochemical characteristics of these films with the
use of a 3-electrode electrochemical cell (see figure 1a).
We have used the techniques of cyclic voltammetry and
spectroelectrochemistry to study the electrochemical
behavior of the films with or without immobilized Cyt-
c. [3] The same setup was also used for the development
of the electrochemical biosensor for H 2O2.
Electrochemical sensing is demonstrated, with a limit of
detection of 20 M (figure 1b). The various detection
Figure 1. (a) 3-electrode electrochemical cell. (b) CVs
obtained for a TiO2-SG/Cyt-c film in a pH 7,
0.01 M PBS, before and after the addition of increasing
amounts (20-320M) of H2O2 at scan rate of 25 mVs-1.
The inset shows the plot of the cathodic peak current
versus the H2O2 concentration obtained from the CV
data.
Conclusions
We conclude that proteins adsorb on the TiO 2 films in a
stable and functional way and the Cyt-c/TiO2 films can
be used for the development of optical and
electrochemical biosensors.
References
1. E. Topoglidis, T. Lutz, J. R. Durrant and E.
Palomares, Bioelectrochemistry 517, 142 (2008).
2. E. Topoglidis, Y. Astuti, F. Duriaux, M. Grtzel and
J. R. Durrant, Langmuir 19, 6894 (2003).
3. S. Katsiaounis, C. Tiflidis and E. Topoglidis,
Electrochemical Characterization of mesoporous
TiO2 films for the development of electrochemical
biosensors, Sensors and Actuators B: Chemical
(2015) in preparation .
Keywords:
Cyt-c immobilization, TiO2 mesoporous films, Degussa
P-25, H2O2 biosensor