Calbindin D-28K is a calcium-binding protein found in the cone but not rod photoreceptor cells in the retinas of a variety of species. Recent studies of the monkey retina indicated that calbindin D-28K may be expressed preferentially in non-foveal regions of the retina. In the current studies of human retinas, immunohistochemical experiments demonstrated that calbindin D-28K is reduced or absent in the fovea and parafovea, but prevalent in the perifovea and periphery. These findings were supported by the quantification of calbindin D-28K in 1-mm trephine punches obtained from different regions of the human retina. The specificity of the anti-calbindin D-28K antibodies used in these studies was confirmed by Western blot analysis using purified calbindin D-28K. The protein was purified from retinal tissue and its identity confirmed by partial amino-acid sequence analysis. The expression of calbindin D-28K did not correlate with the spectral properties of the cones, rather to their position in the retina. The study of spatially expressed genes, like the one encoding calbindin D-28K, may help explain the patterns of retinal degeneration seen in some human cone-rod dystrophies.