Braille readers use a braille display and text-to-speech synthesizer while reading and comprehending mathematical expressions and equations. Teachers need to have technological, pedagogical and content (TPACK) knowledge and skills for exploiting the potential of these devices in mathematics education. They have to understand how the use of assistive technology influences the teaching and learning of mathematics. Therefore, the aim of the current study is to support teachers to better understand the dynamic relation between the different TPACK domains. That is why a professional development course was developed in which five mathematics teachers of braille readers in special secondary education participated. The development of the teachers’ TPACK knowledge was analyzed, and course characteristics that helped to develop this knowledge were identified. The results show an increased awareness of the importance of assistive devices, but only a small positive effect in TPACK knowledge and skills. Course activities related to the braille display, the mathematical braille notation and mathematical vocabulary helped to develop TPACK knowledge. However, course activities related to the text-to-speech synthesizer and working in heterogeneous groups did not work out so well. A better understanding of what teaching mathematics to braille readers means is expected to improve their learning.