Near-infrared fluorophore capped gold nanostars for enhanced detection of choroidal neovascularization in living rabbits using multimodal photoacoustic microscopy, optical coherence tomography, and fluorescence microscopy

Enhanced visualization of choroidal neovascularization (CNV) is critical to the precise diagnosis and treatment for patients with neovascular eye diseases such as wet macular degeneration. Currently, eye clinics lack an imaging modality that can precisely detect and visualize the position and margins of CNV in three-dimensions. This study shows that a unique multimodal photoacoustic microscopy, optical coherence tomography, and florescence microscopy system can accurately identify CNV in living rabbits with high resolution and image contrast at a sub-10-micron scale. In addition, to better visualize CNV and distinguish it from normal blood vessels, the peptide RGD targeting neovascularization was conjugated with gold nanostars (GNS) capped with photostable near-infrared (NIR) fluorophore alexa fluor 790 (AF790). Four New Zealand white rabbits with laser-induced CNV were intravenously administered with 400µL of GNS at a concentration of 5 mg/mL. The accumulation of GNS at CNV was monitored by multimodal OCT, PAM, color fundus photographs, fluorescein angiography (FA), and indocyanine green angiography (ICGA) at various time points: 1 h, 2 h, 4 h, 8 h, 24 h, 48 h, 72 h, day 4, day 7, day 9, day 11, and day 14. The experimental results show that GNS accumulation occurs in the region of CNV. The GNS were detected by all three imaging modalities both in vitro and in living rabbits. The PA signal was increased 19-fold 24 h post-injection. In addition, fluorescence signal gradually decreased over time. Histological analysis and TUNEL assay show no toxicity in the rabbit at the administered concentrations. Therefore, GNS-assisted multimodal imaging has the possibility to improve microvasculature imaging.