Positive and negative affect have been shown to have implications for hormones like cortisol but how moment to moment changes in affect (i.e., affect variability) influence cortisol secretion is less well understood. Additionally, context characteristics such as mean affect and stress may influence the association between affect variability and cortisol output. In the current study, we examined affect, stress, and cortisol data from 113 participants (age range = 25–63, M = 35.63, SD = 11.34; 29% male; 42% White/Caucasian, 37% Asian or Pacific Islander, 13% Hispanic/Latino, 4% Black/African American, 1% Native American, Eskimo, or Aleut, 4% selected “other” for their race/ethnicity). Participants completed ecological momentary assessments assessing positive and negative affect and stress four times per day for five days and provided saliva samples at each time point. Saliva was assayed for cortisol, and area under the curve with respect to ground was computed. In a three-way interaction, both positive affect mean level and stress moderated the association between positive affect variability and cortisol (b = −1.55, t(100) = −3.29, SE = 0.47, p <.01, β = −4.05). When breaking down this three-way interaction, in the context of low stress and high mean positive affect, variability was positively related to total cortisol output. In contrast, in the context of high stress and high mean positive affect, variability was negatively related to total cortisol output. While greater positive affect variability is generally worse for health-relevant outcomes (as prior research has shown and as we show here at low levels of stress), at high levels of stress, fluctuation in affect may be adaptive. For someone experiencing a high stress week, having fluctuations in positive affect may mean that they are adaptively changing to meet their environmental needs especially when they typically report high mean positive affect levels. There were no associations between negative affect variability and cortisol secretion nor did mean negative affect or stress play a moderating role for negative affect variability. This study provides evidence that positive affect variability’s association with cortisol secretion throughout the day may vary based on stress and mean positive affect levels. •For individuals with low stress and high mean positive affect, positive affect variability was positively related to cortisol secretion.•For individuals high stress and high mean positive affect, positive affect variability was negatively related to cortisol secretion.•There were no associations between negative affect variability and cortisol secretion.