This paper aims to determine the prospective biology teachers' self-confidence in technological pedagogical content at Hacettepe University and to compare it with respect to such variables as gender and grade levels. For these purposes, the scale for technological pedagogical content self-confidence, which was originally developed by Graham, Burgoyne, Cantrell and Harris (2009) and was adapted into Turkish by Timur and Tasar (2011), was used. The scale consists of 31 items and is four dimensional. The first dimension is the dimension of Technological Pedagogical Content Knowledge (TPCK), the second is the dimension of Technological Pedagogical Knowledge (TPK), the third is the dimension of Technological Content Knowledge (TCK), and the fourth one is the dimension of Technological Knowledge (TK). Cronbach Alpha reliability coefficient is .92 for the overall scale. The research sample was composed of the reachable 121 of the 134 students studying in the Biology Education department of Hacettepe University. In consequence, it was found that the prospective teachers' self-confidence in terms of technological pedagogical content was high, that there were no differences on gender basis, but that differences were available between the second and fourth year students.