Autonomous motivation for self-set goals (pursuing goals for more intrinsic and less extrinsic reasons) has been linked to affective well-being. Using intensive longitudinal data, the present article examines the link between university students' autonomous study motivation with affective well-being and targets fulfillment of the basic psychological needs as a potential mediating factor of this association on the within-person and the between-person level. University students in Germany (N = 488) completed an online questionnaire once a week over up to two semesters, indicating their weekly study motivation, need fulfillment, and affective well-being. Multilevel structural equation models were employed to target (a) the structure of autonomous study motivation and (b) the associations of autonomous study motivation with need fulfillment and affective well-being. Autonomous study motivation was associated with students' positive and negative affect on both levels. In line with predictions by self-determination theory, multilevel mediation models suggested indirect effects via need fulfillment on both levels. Results highlight the central role of autonomous study motivation and need fulfillment in university students' socioemotional adjustment. A better understanding of this socioemotional adjustment of university students may be an important step for increasing overall study satisfaction and developing interventions to reduce study dropout.