PURPOSE. To image the fundus non-invasively at two different penetration depths using a multispectral imaging system. Monochromatic images at visible spectrum wavelengths and near-infrared wavelengths were qualitatively assessed for choroidal visibility. These images were used calculate oxygen saturation in retinal tissue, optic nerve head tissue, vein, and choroidal tissue in healthy controls and glaucoma patients. METHODS. A fundus camera-based multispectral snapshot oximeter imaged the fundus of healthy subjects and patients with varying ophthalmological pathology. The images of healthy controls and glaucoma patients were analyzed to determine oxygen saturation in the optic nerve head cup and rim, superficial and deep vein, macula and choroidal tissue. RESULTS. Visible: Average oxygen saturation for the ONH cup was 65 Ã‚Â± 6 percent for healthy controls and 61 Ã‚Â± 10 percent for glaucoma patients. For the ONH rim, it was 67 Ã‚Â± 3 percent for healthy controls and 64 Ã‚Â± 17 percent for glaucoma patients. For the vein, it was 67 Ã‚Â± 15 percent for healthy controls and 56 Ã‚Â± 22 percent for glaucoma patients. For the macula, it was 87 Ã‚Â± 10 percent for healthy controls and 93 Ã‚Â± 1 percent for glaucoma patients. NIR: The average oxygen saturation for the vein was 66 Ã‚Â± 20 percent for healthy controls, 58 Ã‚Â± 0.4 percent for glaucoma suspects and 54 Ã‚Â± 17 percent for glaucoma patients. For the choroidal tissue below the macula, it was 99 Ã‚Â± 5 percent in healthy controls and 81 Ã‚Â± 8 percent in glaucoma patients. CONCLUSIONS. Choroidal visibility is enhanced in near infrared monochromatic images from visible spectrum monochromatic images. Oxygen saturation results were lower in glaucoma patients for all anatomical areas analyzed except the avascular macula.