Fabricating protein immunoassay arrays on nitrocellulose using dip-pen lithography techniques

Advancements in lithography methods for printing biomolecules on surfaces are proving to be potentially beneficial for disease screening and biological research. Dip-pen nanolithography (DPN) is a versatile micro and nanofabrication technique that has the ability to produce functional biomolecule arrays. The greatest advantage, with respect to the printing mechanism, is that DPN adheres to the sensitive mild conditions required for biomolecules such as proteins. We have developed an optimised, high-throughput printing technique for fabricating protein arrays using DPN. This study highlights the fabrication of a prostate specific antigen (PSA) immunoassay detectable by fluorescence. Spot sizes are typically no larger than 8 μm in diameter and limits of detection for PSA are comparable with a commercially available ELISA kit. Furthermore, atomic force microscopy (AFM) analysis of the array surface gives great insight into how the nitrocellulose substrate functions to retain protein integrity. This is the first report of protein arrays being printed on nitrocellulose using the DPN technique and the smallest feature size yet to be achieved on this type of surface. This method offers a significant advance in the ability to produce dense protein arrays on nitrocellulose which are suitable for disease screening using standard fluorescence detection.