Quantitative assessment of nerve fiber layer (NFL) thickness in normal and glaucomatous eyes, and correlation with conventional measurements of the optic nerve structure and function. We studied 59 eyes of 33 subjects by conventional ophthalmologic physical examination, Humphrey 24-2 visual fields, stereoscopic optic nerve head photography, and optical coherence tomography. Nerve fiber layer thickness as measured by optical coherence tomography demonstrated a high degree of correlation with functional status of the optic nerve, as measured by visual field examination (P = .0001). Neither cupping of the optic nerve nor neuroretinal rim area were as strongly associated with visual field loss as was NFL thickness (P = .17 and P = .21, respectively). Cupping correlated with NFL thickness only when the cup was small (cup-to-diameter ratio, 0.1 to 0.3) or large (cup-to-diameter ratio, 0.8 to 1.0) (P = .006); there was no correlation between cupping and NFL thickness otherwise. Nerve fiber layer, especially in the inferior quadrant, was significantly thinner in glaucomatous eyes than in normal eyes (P = .04). Finally, we found a decrease in NFL thickness with aging, even when controlling for factors associated with the diagnosis of glaucoma (P = .03). Nerve fiber layer thickness can be measured using optical coherence tomography. These measurements provide good structural and functional correlation with known parameters.