Abstract Teachers’ emotions have been approved to play a pivotal role in higher education. However, the interface of university teachers’ emotions and assessment practices has been widely ignored in second/foreign language contexts. To fill this lacuna, this study examined the perceptions of 35 Iranian EFL university teachers regarding the types, triggers, and regulation strategies of assessment-related emotions through a semi-structured interview. After a thematic analysis of the data by MAXQDA software, it was found that Iranian university teachers have experienced both negative and positive emotions during their assessment practices. Such emotions were mainly triggered by teachers’ assessment methods/practices, teaching context, and the assessment culture of the department. Furthermore, the results indicated that the participants employed several preventive and responsive strategies to regulate negative and positive feelings during the L2 assessment. Practical implications are discussed regarding the necessity of training university teachers on the interplay of emotions and L2 assessment.