Starch-gel electrophoresis was used to assess temporal and spatial genetic variation in populations of white-tailed deer (Odocoileus virginianus) in western Tennessee. Samples of liver and kidney obtained from animals at five localities during 1985–1992 were analyzed at 11 loci known to be polymorphic in white-tailed deer. There were minimal significant differences in allelic frequencies between sexes and among age groups for each year at each locality and among years at each locality with ages and sexes combined. No significant difference occurred among mean values of heterozygosity. A heterozygote deficiency was detected in white-tailed deer populations from western Tennessee and could reflect limited inbreeding but was more likely due to Wahlund effects. Limited temporal variation occurred among years within a locality and indicated a stasis in terms of minimal genetic drift and harvest effects on genetic structure of the populations. Herd origin, stocking history of populations, and gene flow were probable causes of spatial heterogeneity in deer populations in western Tennessee.