Team-based learning (TBL) stresses applying knowledge rather than absorbing knowledge in class; studies have investigated the use of TBL and its merits in different teaching courses (e.g., medical science and business). TBL is most effective when students learn autonomously before class. However, the ability of autonomous learning is highly associated with the ability of self-regulated learning (SRL); most importantly, not every student possesses good (or high) SRL ability. Nevertheless, few studies have compared the effectiveness of TBL in students with different SRL abilities. To address this issue, this study analyzed approximately 90 students, whose course teaching involves office application software (Microsoft Excel). This study also developed an online TBL system (called Online TBL) to facilitate performing TBL and to collect the learning behaviours of students with different (high or low) SRL abilities on each TBL stage. The analytical results show that compared with the low-SRL students, the high-SRL students were more prepared for class because they spent more reviewing material and had better scores on personal uploaded Excel and Individual Readiness Assurance Test. From the feedback of the peer evaluation, the results also show that the high-SRL students received more credits than the low-SRL students did. The questionnaire survey revealed that both low-SRL and high-SRL students had a favourable impression of TBL. Further discussion is given to explain the above results.