This article analyses the challenging collaboration between small and medium-sized enterprises (SMEs) and knowledge institutions. The aim of the article is to shed light on the barriers, which hinder collaboration, and to investigate the particular role of research and technology organizations (RTOs) as potential mediators of collaboration between SMEs and universities. On the basis of a unique sample consisting of 151 SMEs, RTOs and universities from seven countries, the differences across dyads of potential collaborations are identified. In particular, the article finds that both firms and universities with collaboration experience with the other partner in general perceive higher barriers than inexperienced firms or universities. In terms of the mediating role of RTOs, the article illustrates that universities perceive lower barriers when collaborating with RTOs than with SMEs. A similar tendency to a mediating role of RTOs can be found among the SMEs' perception of university collaboration. Finally, the analysis shows that the knowledge institutions perceive the SMEs as very important collaboration partners, but the same sense of importance is not shared by the SMEs regarding collaboration with the knowledge institutions. Hence, there seems to be a quite unbalanced view on the potential of the collaborative ties, which is further discussed in the conclusion.