Immobilisation of oligo-peptidic probes for microarray implementation : Characterisation by FTIR, Atomic Force Microscopy and 2D fluorescence

• Proteomic microarrays show a wide range of applications for the investigation of DNA-protein, enzyme-substrate as well as protein-protein interactions. Among many challenges to build a viable protein microarray, the surface chemistry that will allow to immobilised various proteins to retain their biological activity is of paramount importance. Here we report a chemical functionalisation method allowing immobilisation of oligo-peptides onto silica surface (porous silica, glass, thermal silicon dioxide). Substrates were first derivatised with a monofunctional silane allowing the elaboration of dense and uniform monolayers in highly reproducible way. Prior to the oligo-peptides grafting, this organic layer was functionalised with an amino-polyethyleneglycol. The coupling step of oligo-peptides onto functionalised supports is achieved through activation of the C-terminal function of the oligo-peptides. Chemical surface modifications were followed by FTIR spectroscopy, AFM measurements and fluorescence scanning microscopy. A systematic study of the oligo-peptide grafting conditions (time, concentration, solvent) was carried out to optimise this step. The oligo-peptides grafting strategy implemented in this work ensure a covalent and oriented grafting of the oligo-peptides. This orientation is ensured through the use of fully protected peptide except the terminal primary an-tine. The immobilized peptides will be then deprotected before biological recognition. This strategy is crucial to retain the biological activity of thousands of oligo-probes assessed on a microarray.

Nyckelord

oligo-peptide immobilization

microarray

biotin

streptavidin

FTIR

AFM

fluorescence

protein microarrays

technology

chip

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