Gold surface functionalization and patterning for specific immobilization of olfactory receptors carried by nanosomes.
There is substantial interest in engineering solid supports to achieve functional immobilization of membrane receptors both for investigation of their biological function and for the development of novel biosensors. Three simple and practical strategies for immobilization of a human olfactory receptor carried by nanosomes are presented. The basis of the functionalization of solid gold surfaces is a self-assembled monolayer (SAM) containing biotinyl groups. Biotinyl groups are subsequently used to attach neutravidin and then biotinylated monoclonal antibody directed against the receptor to allow its specific grafting. Surface plasmon resonance technique is implemented for real-time monitoring of step-by-step surface functionalization and, in addition, for testing the functional response of immobilized olfactory receptors. We show that OR1740 is functional when immobilized via a tag attached to its C-terminus, but not via its N-terminus. Finally, we demonstrate that gold surfaces can be patterned by the SAMs tested using microcontact printing. AFM images of immobilized nanosomes onto a patterned surface suggest that small nanosomes flatten and fuse into larger vesicles but do not merge into a continuous layer. The whole study emphasizes the outstanding performances of the BAT/PEGAT SAM, which could be useful for developing on-a-chip sensor formats for membrane receptor investigations and use.
Vidic
Jasmina
J
Neurobiologie de l'Olfaction et de la Prise Alimentaire, Equipe Récepteurs et Communication Chimique, INRA, Jouy en Josas, France. jasmina.vidic@jouy.inra.fr
Pla-Roca
Mateu
M
Grosclaude
Jeanne
J
Persuy
Marie-Annick
MA
Monnerie
Régine
R
Caballero
David
D
Errachid
Abdelhamid
A
Hou
Yanxia
Y
Jaffrezic-Renault
Nicole
N
Salesse
Roland
R
Pajot-Augy
Edith
E
Samitier
Josep
J
D000911:Antibodies, Monoclonal / Q000737:chemistry; D001360:Avidin / Q000737:chemistry; D015374:Biosensing Techniques / Q000379:methods*; D006046:Gold / Q000737:chemistry*; D006801:Humans; D008567:Membranes, Artificial; D018625:Microscopy, Atomic Force / Q000379:methods; D053758:Nanoparticles / Q000737:chemistry* / Q000648:ultrastructure; D010316:Particle Size; D018035:Receptors, Odorant / Q000737:chemistry*; D012680:Sensitivity and Specificity; D020349:Surface Plasmon Resonance / Q000379:methods; D013499:Surface Properties; D013997:Time Factors
D000911:Antibodies, Monoclonal; D008567:Membranes, Artificial; D018035:Receptors, Odorant; C093483:neutravidin; D001360:Avidin; D006046:Gold