The current study investigates the beneficial combination of optical coherence tomography (OCT) and photoacoustic microscopy (PAM) as a safe method for observing retinal and choroidal vasculature. A recent addition to the field has been the integration of gold nanoparticles (AuNPs) to provide enhanced contrast in OCT and PAM images. The improved analysis of capillaries is the result of the strong optical scattering and optical absorption of gold nanoparticles due to surface plasmon resonance. Femtosecond laser ablation created the ultra-pure colloidal gold nanoparticles, which were then capped with polyethylene glycol (PEG). The AuNPs were administered to thirteen New Zealand rabbits to determine the advantages of this technology, while also investigating the safety and biocompatibility. The study determines that the synthesized PEG-AuNPs (20.0 ± 1.5 nm) were beneficial in enhancing contrast in PAM and OCT images without demonstrating cytotoxic effects to bovine retinal endothelial cells. In living rabbits, the administered PEG-AuNPs resulted in an 82% increased signal for PAM and a 45% increased signal for OCT in the retinal and choroidal vessels. A histology and biodistribution report determined that the AuNPs had mostly accumulated in the liver and spleen. TUNEL staining and histology established that no cell injury or death in the lung, liver, kidney, spleen, heart, or eyes had occurred up to 1 week after receiving a dose of AuNP. The nanoparticle technology, therefore, provides an effective and safe method to enhance contrast in ocular imaging, resulting in improved visualization of retinal microvasculature.