PURPOSE Although the effect of spatial sampling on the visibility of grating stimuli is well described and well understood, little research has been conducted into the effects of spatial sampling on the visibility of letter optotypes. The purpose of this study was to investigate whether thresholds for spatially sampled Sloan letters were equal to or significantly smaller than the spacing between spatial samples. METHODS For four visually normal subjects, we measured Sloan letter acuity thresholds, presented on a computer monitor, after the letters had been sampled by sampling arrays with 6.285 elements per square degree, (23.9 min of arc between samples for a square packed array). We used four different sampling arrays: square packed; hexagonally packed; a cone-like array with positive contrast; and a cone-like array with negative contrast. Thresholds were assessed using letter-counting rules and also Probit analysis. RESULTS Although results depended on array type, and the definition of sample spacing, letter acuity thresholds were substantially less than estimates of sample spacing by between 0.290 log units (49% less) (hexagonally sampled Probit thresholds compared with spacing between hexagonal samples) to 0.136 log units (27% less) (positive contrast cone-like sampled letter-counting thresholds compared with spacing between rows of hexagonal samples). CONCLUSIONS Sample spacing is not an absolute limit for Sloan letter thresholds. By comparison with previous measurements of human foveal cone sampling of space, our findings suggest that cone sampling limits for Sloan letters could be as small as 20/4.