In this work, a collimated helium beam was used to activate a thiol-poly(ethylene glycol) (SH-PEG) monolayer on gold to selectively capture proteins in the exposed regions. Protein patterns were formed at high throughput by exposing a stencil mask placed in proximity to the PEG-coated surface to a broad beam of helium particles, followed by incubation in a protein solution. Attenuated Total Reflectance-Fourier Transform Infrared Spectroscopy (ATR-FTIR) spectra showed that SH-PEG molecules remain attached to gold after exposure to beam doses of 1.5-60 μC/cm2 and incubation in PBS buffer for one hour, as evidenced by the presence of characteristic ether and methoxy peaks at 1120 cm-1 and 2870 cm-1, respectively. X-ray Photoelectron Spectroscopy (XPS) spectra showed that increasing beam doses destroy ether (C-O) bonds in PEG molecules as evidenced by the decrease in carbon C1s peak at 286.6 eV and increased alkyl (C-C) signal at 284.6 eV. XPS spectra also demonstrated protein capture on beam-exposed PEG regions through the appearance of a nitrogen N1s peak at 400 eV and carbon C1s peak at 288 eV binding energies, while the unexposed PEG areas remained protein-free. The characteristic activities of avidin and horseradish peroxidase were preserved after attachment on beam-exposed regions. Protein patterns created using a 35 μm mesh mask were visualized by localized formation of insoluble diformazan precipitates by alkaline phosphatase conversion of its substrate bromochloroindoyl phosphate-nitroblue tetrazolium (BCIP-NBT) and by avidin binding of biotinylated antibodies conjugated on 100 nm gold nanoparticles (AuNP). Patterns created using a mask with smaller 300 nm openings were detected by specific binding of 40 nm AuNP probes and by localized HRP-mediated deposition of silver nanoparticles. Corresponding BSA-passivated negative controls showed very few bound AuNP probes and little to no enzymatic formation of diformazan precipitates or silver nanoparticles.
ASJC Scopus subject areas
- Biochemistry, Genetics and Molecular Biology(all)
- Agricultural and Biological Sciences(all)