This paper describes studies of surface-patterned nanohydrogels and their use as possible substrates for high-density and high-sensitivity protein arrays. Nanohydrogels, approximately 200 nm in diameter, were created by locally crosslinking dry amine-terminated poly(ethylene glycol) [PEG] (5000 Da) thin films using a focused electron beam. These gels then had a dry height of 50 nm and a swell ratio of about five. They were patterned into arrays with approximately 1 μm inter-gel spacing. These arrayed gels were functionalized with Zinc Finger 9 (ZNF9), a nucleic-acid binding protein. As part of ongoing research, we are interested in how the performance of the functionalized gels compares to that of current commercial substrates. We show, using an assay that binds α-GST antibody to mediate attachment of GST-tagged ZNF9, that nanohydrogels have a consistently higher combination of fluorescent signal and signal-to-noise (SNR) ratio than the comparable commercial substrates. We further show that the SNR characteristic of our assay degrades slower on the nanohydrogel array than on a comparable microarray printed on an epoxide substrate, and we speculate that the difference is in part attributable to the fact that the nanohydrogel array presents a softer and more hydrophilic surface.