Affective responses to physical activity are assumed to play a role in exercise initiation and maintenance. The Physical Activity Affect Scale measures four dimensions of an individual's affective response to exercise. Group differences in the interpretation of scale items can impact the interpretability of mean differences, underscoring the need to examine whether measurement structure holds across groups (e.g., active vs. non-active). In the current study, physically active (n = 158) and non-active (n = 179) participants exercised for 30 min; affect was assessed during exercise (i.e., 20 min). Tests of measurement invariance were conducted in a confirmatory factor analytic framework. There was good evidence for measurement invariance of the Physical Activity Affect Scale across active and non-active groups, with a single non-invariant factor loading, invariance in item intercepts, and invariance in item uniquenesses, although patterns of factor covariances and means differed across groups. Recommendations and implications of research are provided. (Contains 1 table, 1 figure, and 1 footnote.)