To assess individual students’ abilities and misconceptions in mathematics, teachers need diagnostic competencies. Although research has addressed the quality of teachers’ diagnostic competencies in recent years, it is not very clear how to foster these competencies effectively in the course of prospective teachers’ university education. Research suggests that simulations with instructional support are promising tools for fostering complex competencies. We have developed a simulation that aims at measuring and fostering prospective primary school teachers’ competencies to assess students’ mathematical abilities and misconceptions based on their written task solutions. In this study, we analysed data from prospective primary school mathematics teachers who used one of three different versions of the simulation. Two versions contained a specific type of scaffolding, while the third version did not contain scaffolding. Specifically, the two scaffolding types were content-related scaffolding that emphasized the use of specific pedagogical content knowledge, and strategic scaffolding that emphasized diagnostic activities. The results suggest that integrating scaffolding into the simulation did not substantially influence participants’ overall perception of the simulation regarding presence, authenticity, or perceived cognitive load. Compared to participants in a control group without intervention, participants who used the simulation with scaffolding had higher diagnostic accuracy regarding overall assessment of students’ competence level. However, only content-related scaffolding but not strategic scaffolding or no scaffolding tended to improve participants’ competence in identifying students’ specific misconceptions. The results provide a first empirical basis for further development of the simulation.