The purpose of this study was to determine whether ultrastructural changes in intramyocellular lipid (IMCL) and mitochondria occur with aging. Muscle samples were analyzed from 24 young and 20 old, equally active, individuals for IMCL and mitochondria quantity and size as well as their association. Old men had larger IMCL droplets than all other groups in the total muscle area. Old individuals showed higher IMCL content in the subsarcolemmal area. Young participants had a greater number of mitochondria compared with old participants in both fiber regions and greater enzyme activities of cytochrome c oxidase and citrate synthase. The fraction of IMCL touching mitochondria was lowest in old women in the total area and in old men in the subsarcolemmal region. In summary, older adults have larger IMCL droplets, fewer mitochondria, and a lower proportion of IMCL in contact with mitochondria. These factors likely contribute to age-related reductions in mitochondrial function and lipid metabolism.