Despite the interest in mindfulness over the past 20 years, studies have only recently begun to examine mindfulness in older adults. The primary aim of this study was to evaluate pretreatment to post-treatment change in negative affect variability (NAV) following a mindfulness training among 134 mildly stressed, middle-aged to older adults. The secondary aim was to assess if the effects of mindfulness training on NAV would be partially explained by pretreatment to post-treatment reductions in perceived stress, a trend that would be congruent with several stress models. In this randomized control trial, participants were assigned to either a 6-week mindfulness meditation training programme or to a wait list control. Ecological momentary assessment, a data capturing technique that queries about present moment experiences in real time, captured NAV. Mixed-model ANOVAs and a path analysis were conducted. Participants in the mindfulness meditation training significantly reduced NAV when compared with wait list control participants. Further, there was a significant indirect group effect on reductions in NAV through change in perceived stress. Few studies have tested mechanisms of action, which connect changes that occur during mindfulness training with psychological outcomes in older adults. Understanding the mechanisms by which mindfulness enhances well-being may optimize interventions.