Although a number of parameters have been standardized when testing visual acuity (including chart distance, optotypes and luminance of the chart), there is considerable variation in room lighting conditions used. Currently no research exists which either suggests a particular room illumination, or even if there is any difference in visual acuity with different room lighting conditions. The purpose of the current study was to determine whether changes in room illumination affect the level of distance visual acuity recorded. Visual acuity was randomly tested on 50 subjects (98 eyes) using the standard Snellens chart in two different room illumination levels, with normal room illumination (with room lights on- 1300 lux) and with reduced room illumination (with room lights off – 90 lux). Residual refractive error (difference between spectacle correction and autorefraction) was calculated and pupil size measured in each condition. Overall a significant difference in visual acuity between the two lighting conditions was found with visual acuity levels improving with room illumination (6/9 + 4 in illuminated room, 6/9 +2 in non-illuminated room; t=4.653, p<0.001). The difference was found to be greater in the non-emmetropic group (6/12+3 in illuminated room, 6/12 in non-illuminated room). There was a small subgroup of subjects’ eyes where the visual acuity level dropped by more than one line in the non-illuminated room (n=18). The reason for this difference may be related to optical influences on visual acuity such as accommodation and night myopia.