In vivo ZW800-microbead imaging of retinal and choroidal vascular leakage in mice

The eye is an attractive organ for non-invasive discovery and monitoring of disease progression. Traditionally, fluorescein angiography (FA) and indocyanine green angiography (ICGA) have been used for dynamic evaluation of the retina and its vasculature. However, both fluorescein and indocyanine green (ICG) possess considerable disadvantages. FA is limited to assessing superficial retinal blood flow and often results in an unclear view due to fluorescein leakage. This obscures important pathologies such as neovascularization, ischemia and inflammation. ICG, a near-infrared fluorophore (NIRF), has nonspecific binding, high uptake and retention in tissues, as well as detrimental effects on the hepatobiliary tract. Here, we present a potential contrast agent for imaging ocular vascular permeability with ZW800, a heptamethine indocyanine NIRF, conjugated to polystyrene latex beads (ZW800m). ZW800 is an excellent alternative for near-infrared imaging, as it has excellent contrast, superior clearance, and is amendable to conjugation. ZW800m conjugation is an easy, attractive method of in vivo imaging and real-time tracking of ocular vascular pathologies. ZW800m is readily imaged via commercially available laser ophthalmoscope (SLO, HRA OCT, Spectralis) to assess vascular permeability in the mouse retina and choroid. In Type 1 diabetic Ins2Akita mice, ZW800m was observed in mouse retina but not in wild-type mice. After laser-induced choroidal neovascularization (CNV), ZW800m was observed in mouse choroid but not in control. In both CNV and diabetic mice, ZW800 imaging showed increased hyperfluorescence on ICG modality (ICGA) not seen on FA. Presence of ZW800m in respective tissues was confirmed ex vivo with flatmounts visualized with EVOS 800 nm light cube. ZW800 imaging may be easily employed in the research laboratory.