ADIBO-based "click" chemistry for diagnostic peptide micro-array fabrication: physicochemical and assay characteristics

• Published in: Molecules (Basel, Switzerland) Vol. 18; no. 8; pp. 9833 - 9849
• Main Authors: Prim, Denis, Rebeaud, Fabien, Cosandey, Vincent, Marti, Roger, Passeraub, Philippe, Pfeifer, Marc E
• Format: Journal Article
• Language: English
• Published: Switzerland MDPI AG 16-08-2013; MDPI
• Subjects: ADIBO; Antibodies; antibody diagnostics; Automation; Azide; blocking reagents; Chemistry; Click Chemistry - methods; Copper; Diagnostic tests; Enzyme-Linked Immunosorbent Assay; Immunoassay - methods; Kinetics; non-specific binding (NSB); Oxidation; peptide microarray; Peptides; Peptides - chemistry; Polyethylene glycol; Reagents; SPAAC; Switzerland
• ISSN: 1420-3049; 1420-3049
• DOI: 10.3390/molecules18089833

Several azide-derivatized and fluorescently-labeled peptides were immobilized on azadibenzocyclooctyne (ADIBO)-activated slide surfaces via a strain-promoted alkyne-azide cycloaddition (SPAAC) reaction revealing excellent immobilization kinetics, good spot homogeneities and reproducible fluorescence signal intensities. A myc-peptide micro-array immunoassay showed an antibody limit-of-detection (LOD) superior to a microtiter plate-based ELISA. Bovine serum albumin (BSA) and dextran covalently attached via "click" chemistry more efficiently reduced non-specific binding (NSB) of fluorescently-labeled IgG to the microarray surface in comparison to immobilized hexanoic acid and various types of polyethylene glycol (PEG) derivatives. Confirmation of these findings via further studies with other proteins and serum components could open up new possibilities for human sample and microarray platform-based molecular diagnostic tests.