Surface immobilisation of antibody on cyclic olefin copolymer for sandwich immunoassay.
In this work, the surface functionalisation of the commercially available cyclic olefin copolymer (COC) materials, Zeonor and Zeonex, has been studied. The methodology employed involved oxidation in oxygen plasma, functionalisation of the oxidized surface with aminopropyl triethoxy silane and, finally, attachment of antibody using covalent linker molecules. 1,4-Phenylene diisothiocyanate was selected as the most suitable cross-linker for the attachment of protein, as assessed by fluorescent intensity measurements on immobilised FITC-labelled IgG antibody. The modification method was characterised by contact angle measurements, ellipsometry, X-ray photoelectron spectroscopy (XPS) and fluorescence microscopy. The data are consistent with the deposition of a polymeric film of the silane chemisorbed to the oxidised plastic surface. The functionalised surfaces were employed in a sandwich immunoassay format using the reagents goat anti-human IgG (G alphaHIgG) and fluorescently labelled G alphaHIgG (Cy5-G alphaHIgG) as capture and detection antibodies, respectively, and with human IgG (HIgG) as the model analyte. The lowest concentration of HIgG detected was 0.1 ng ml(-1), with a relative standard deviation of 15%. Non-specific binding effects were also assessed. The method and supporting data demonstrate that simple approaches to surface functionalisation can be adapted to plastic-based devices.
Raj
Jog
J
Biomedical Diagnostics Institute Programme, Tyndall National Institute, Lee Maltings, University College, Lee Maltings, Cork, Ireland.
Herzog
Grégoire
G
Manning
Mary
M
Volcke
Cédric
C
MacCraith
Brian D
BD
Ballantyne
Scott
S
Thompson
Michael
M
Arrigan
Damien W M
DW
D000327:Adsorption; D000906:Antibodies / Q000737:chemistry* / Q000276:immunology; D015374:Biosensing Techniques / Q000295:instrumentation*; D003516:Cycloparaffins / Q000737:chemistry*; D004867:Equipment Design; D019544:Equipment Failure Analysis; D006801:Humans; D007118:Immunoassay / Q000295:instrumentation*; D007074:Immunoglobulin G / Q000032:analysis* / Q000276:immunology; D011485:Protein Binding; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D000906:Antibodies; D003516:Cycloparaffins; D007074:Immunoglobulin G