Biosensors for the detection and monitoring of ... - Archimer - Ifremer

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19 Φεβ 2013 (πριν από 4 χρόνια και 3 μήνες)

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Biosensors for the detection and monitoring of
Alexandrium minutum along the French coasts.
1 2 3 2 2 2 3
E. Antoine , C. Compère , S. Cosnier , M.P. Crassous , H. Derbal , E. Erard-Le Denn , C. Gondran ,
2 2 2 4 5 6 6
E. Guilminot , M. Lehaitre , F. Mazéas , H. Moreau , H. Perrot , C.M. Pradier , M. Salmain
CENTRE NATIONAL
DE LA RECHERCHE
SCIENTIFIQUE
Introduction
Maximum
concentration per site
The microalgae Alexandrium minutum produces neurotoxins responsible for
Cells/l
paralytic shellfish poisoning, which upon accumulation in shellfish represent a
human health risk. This species recurrently forms toxic blooms in the Northern
6
10
part of the French Brittany coast. At present, algae identification relies on
6
to 10
tedious microscope observations mostly performed by skilled taxonomists in
laboratories. Newly emerging fields of molecular taxonomy, nanoscale
technology and biotechnology offer opportunities for the development of in-
REPHY monitoring network along the French coasts
situ biosensors for the detection and the monitoring of toxic algae.
Maximum of Alexandrium cells/l between 1999-2001
http://www.ifremer.fr/envlit/surveillance/rephy.htm
Monoclonal antibody specific to
Strategy
Alexandrium species: Hybridoma 12a10
Biotinylated polypyrrole modified
Transducers: Au, Pt, stainless steels
from Arago laboratory
quartz quartz-crystal microbalance
[Detal, Rapid detection of toxic algae,
S. Cosnier, H. Perrot et R. Wesssel, (2001)
Ref n°QLRT-1999-30778]
Electroanalysis, 13, 11 971.
Cell surface
monoclonal
Gravimetric or
SEM-micrograph of
Amplification
Signal processing
antibody or
electrical or
A. minutum (17-25 µm)
optical transducer
Nucleic acid
probe


Immobilisation of bioreceptors via
Light micrograph of A. minutum
Hybridization on supports treated with
- self-assembled monolayers (Au surface)
[C-M. Yam, C.M. Pradier, M. Salmain, P. Marcus, G. Jaouen avidin, revelation by a green coloured
(2001) J. of Colloid and Interface Science, vol. 235, 183-189]
reaction of the ABTS substrate
Amin2 nucleic probe for Alexandrium minutum
- electropolymerized biotinylated polypyrrole
A : Positive reference,
5'< AGC ACT GAT GTG TAA GGG CT <3' films (Pt surface)
B : Functionalized Pt surface
[Guillou, L. et al.(2002) Protist, Vol. 153, 223-238] [S. Cosnier (1999) Biosensors and Bioelectronics Vol.14,443]
C : Functionalized Au surface
The characterization of the biomolecules immobilisation is extremely significant to improve the stability and the response of the biosensor
1,4
1116
1635
Nature of the
1704
1481
3590
molecular groups
b
a
and their
0,8
orientation from
4000 3500 3000 2500 2000 1500 1000
wavenumber (cm-1)
FT-IRRAS
FT-IRRAS spectrum on functionalized surfaces: a/ Au, b/ Pt
Conclusions
Due to the complexity and to the interdisciplinary nature of such a development, this research requires a
collaboration between fields of material science, marine chemistry, microbiology, biochemistry, biology and
marine technology. The final goal will be the development of an in situ instrumentation for rapid, sensitive and
selective in situ detection of toxic blooms in the frame of coastal waters monitoring.
1
Ifremer, Centre de Nantes, rue de l'Ile d'Yeu, BP 21105, 44311 Nantes Cedex 3, France
2
Ifremer, Centre de Brest, B.P. 70, 29280 Plouzané, France
3
Laboratoire d'Electrochimie Organique et de Photochimie Redox, Bâtiment Chimie Recherche, Université Joseph Fourier Grenoble 1, 301 rue de la Chimie, BP 53, 38041 Grenoble Cedex 9, France.
4
Laboratoire Arago, Université Paris VI “ Modèles en biologie cellulaire et évolutive ”, BP 44, 66651 Banyuls sur mer, France.
5
UPR 15 du CNRS, Laboratoire de Physique des Liquides et Electrochimie du CNRS, Université P. et M. Curie,Tour 22, 5ème étage, 4, place Jussieu, 75252 Paris cedex 05, France.
6
Ecole Nationale Supérieure de Chimie de Paris - Laboratoire de Physico-Chimie des Surfaces, 11, rue Pierre et Marie Curie, 75005 Paris, France.
Normalised intensity (a. u.)