In quadrupeds, the three-dimensional orientation of the heart with respect to the thorax is fundamentally different from that in humans. In this study, we assessed the best surgical approach to the tricuspid valve in sheep. Firstly, different surgical access sites to the tricuspid valve were tested in sheep cadavers, the anatomy was analyzed, and the optimal surgical approach to the tricuspid valve was determined. Secondly - along with cardiopulmonary bypass and cardioplegic arrest -the chosen approach was tested in six adult sheep in vivo. Anatomical analyses revealed that a left thoracotomy provided optimal access to the aorta and left heart. However, visualization of the right heart was significantly impaired. In contrast, a right thoracotomy provided good access to the right heart, but the ascending aorta was difficult to approach. Therefore, in the in vivo studies, arterial cannulation was performed through a carotid (n = 4) or femoral (n = 2) artery. In conclusion, a right-sided thoracotomy allows good visualization of all components of the tricuspid valve complex in sheep, but not of the ascending aorta. Consequently, peripheral vessels are preferred for arterial cannulation. This work may stimulate the investigation of pathomechanisms and/or novel treatment options for tricuspid valve pathologies.