Immobilisation of oligo-peptidic probes for microarray implementation: Characterisation by FTIR, Atomic Force Microscopy and 2D fluorescence
2005 (English)In: Journal of chromatography. B, ISSN 1570-0232, E-ISSN 1873-376X, Vol. 822, no 02-jan, 304-310 p.Article in journal (Refereed) Published
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.
Place, publisher, year, edition, pages
2005. Vol. 822, no 02-jan, 304-310 p.
oligo-peptide immobilization, microarray, biotin, streptavidin, FTIR, AFM, fluorescence, protein microarrays, technology, chip
IdentifiersURN: urn:nbn:se:kth:diva-14961ISI: 000231008900039OAI: oai:DiVA.org:kth-14961DiVA: diva2:333002
QC 201005252010-08-052010-08-05Bibliographically approved