Immobilization of rhodopsin on a self-assembled multilayer and its specific detection by electrochemical impedance spectroscopy.
Rhodopsin, the G protein-coupled receptor (GPCR) which mediates the sense of vision, was prepared from calf eyes and used as receptor enriched membrane fraction. In this study it was immobilized onto gold electrode by two different techniques: Langmuir-Blodgett (LB) and a strategy based on a self-assembled multilayer. We demonstrated that Langmuir and LB films of rhodopsin are not stable. Thus, in this study a new protein multilayer was prepared on gold electrode by building up layer-by-layer a self-assembled multilayer. It is composed of a mixed self-assembled monolayer formed by MHDA and biotinyl-PE, followed by a biotin-avidin system which allows binding of biotinylated antibody specific to rhodopsin. The immobilization of rhodopsin in membrane fraction, by the specific antibody bound previously on self-assembled multilayer, was monitored with electrochemical impedance spectroscopy (EIS). In addition, the specificity and sensitivity of this self-assembled multilayer system to the presence of rhodopsin were investigated. No effect was observed when the system was in contact with olfactory receptor I7 in membrane fraction used for control measurements. All these results demonstrate that rhodopsin can be immobilized efficiently, specifically, quantitatively and stably on gold electrode through the self-assembled multilayer.
Hou
Yanxia
Y
Centre de Génie Electrique de Lyon (CEGELY), Ecole Centrale de Lyon, B.P.163, 69134 Ecully Cedex, France.
Helali
Salwa
S
Zhang
Aidong
A
Jaffrezic-Renault
Nicole
N
Martelet
Claude
C
Minic
Jasmina
J
Gorojankina
Tatiana
T
Persuy
Marie-Annick
MA
Pajot-Augy
Edith
E
Salesse
Roland
R
Bessueille
Francois
F
Samitier
Josep
J
Errachid
Abdelhamid
A
Akimov
Vladimir
V
Reggiani
Lino
L
Pennetta
Cecilia
C
Alfinito
Eleonora
E
D000327:Adsorption; D015374:Biosensing Techniques / Q000295:instrumentation / Q000379:methods*; D020099:Coated Materials, Biocompatible / Q000032:analysis* / Q000737:chemistry*; D003460:Crystallization / Q000379:methods; D017097:Electric Impedance; D004563:Electrochemistry / Q000379:methods*; D008567:Membranes, Artificial; D011485:Protein Binding; D012243:Rhodopsin / Q000032:analysis* / Q000737:chemistry* / Q000648:ultrastructure; D013057:Spectrum Analysis / Q000379:methods*; D013499:Surface Properties
D020099:Coated Materials, Biocompatible; D008567:Membranes, Artificial; D012243:Rhodopsin