An oxime-based glycocluster microarray.

Carbohydrate microarrays represent powerful tools to study and detect carbohydrate-binding proteins, pathogens or cells. In this paper, we report two original oxime-based methods to prepare surfaces displaying well-defined structures and valency in a given microspot with improved recognition potency with lectins. In a first "direct" approach, fully synthetic aminooxylated glycoclusters have been coated onto aldehyde-activated SiO2 (silicium substrate doped with 50 nm thermal oxide layer). To improve the preparation of the microarray in terms of rapidity and simplicity and to provide addressable surfaces on which sugars can be linked chemoselectively as clusters at defined plots, a second "indirect" strategy has been developed using successive oxime ligation steps. In both cases, binding assays with labelled lectins have revealed more potent and selective interaction due to the clustered presentation of sugars. The observed differences of interaction have been confirmed in solution by ITC.

Surface patterning of (bio)molecules onto the inner wall of fused-silica capillary tubes.

An efficient photochemical method for the site-specific immobilization and patterning of (bio)molecules inside glass capillary tubes is reported. The strategy involves the photodeprotection of reactive aminooxy groups on surfaces and subsequent reaction with aldehyde containing (bio)molecules.

A novel heterotrifunctional peptide-based cross-linking reagent for facile access to bioconjugates. Applications to peptide fluorescent labelling and immobilisation.

A convenient, versatile and straightforward synthesis of a novel heterotrifunctional peptide-based linker molecule is described. This generic bio-labelling reagent contains an amine-reactive N-hydroxysuccinimidyl carbamate moiety, an aldehyde/ketone-reactive aminooxy group and a thiol group with a propensity to form urea, oxime and thioether linkages respectively. The full chemical orthogonality between the free aminooxy and thiol functionalities was demonstrated through the preparation of a fluorescent reagent suitable for the selective staining of a carboxaldehyde-modified surface by means of oxime ligation. The absence of reactivity of these two functions toward the nucleophile-sensitive active carbamate was obtained by using temporary aminooxy- and thiol-protecting groups removable under mild conditions. The utility of the linker molecule to cross-link three different molecular partners has been illustrated by the preparation of fluorescent tripod-functionalised surfaces which may be useful in developing new peptide microarrays and related immunosensors.

Use of gamma-aminopropyl-coated glass surface for the patterning of oligonucleotides through oxime bond formation.

The present work reports on the preparation of glass surfaces coated with NPPOC-protected aminooxy groups and their use for the patterning of oligonucleotides on glass slides and in capillary tubes. The method involves the use of surfaces coated with amino groups using (gamma-aminopropyl)triethoxy silane and subsequent grafting of the aminooxy groups by using the activated ester 1. The NPPOC-protected aminooxy groups on the surfaces can be cleaved upon irradiation. The free aminooxy groups so obtained are subsequently reacted with aldehyde-containing oligonucleotides to achieve efficient surface patterning.

Efficient surface patterning of oligonucleotides inside a glass capillary through oxime bond formation.

The efficient surface patterning of oligonucleotides was accomplished onto the inner wall of fused-silica capillary tubes as well as on the surface of glass slides through oxime bond formation. The robustness of the method was demonstrated by achieving the surface immobilization of up to three different oligonucleotide sequences inside the same capillary tube. The method involves the preparation of surfaces grafted with reactive aminooxy functionalities masked with the photocleavable protecting group, 2-(2-nitrophenyl) propyloxycarbonyl group (NPPOC). Briefly, NPPOC-aminooxy silane 1 was prepared and used to silanize the glass surfaces. The NPPOC group was cleaved under brief irradiation to unmask the reactive aminooxy group on surfaces. These reactive aminooxy groups were allowed to react with aldehyde-containing oligonucleotides to achieve an efficient surface immobilization. The advantage associated with the present approach is that it combines the high-coupling efficiency of oxime bond formation with the convenience associated with the use of photolabile groups. The present strategy thus offers an alternative approach for the immobilization of biomolecules in the microchannels of "labs on a chip" devices.

Oxime bond formation for the covalent attachment of oligonucleotides on glass support.

The chemical attachment of oligonucleotides on glass slides has been achieved using oxime bond formation. This method has been shown very efficient by comparison with the attachment of amino-oligonucleotides via reductive amination.