Objective To summarize the anatomic features of the posterior septum of the knee joint and its application in posterior trans-septal portal for arthroscopic surgery. Methods The literature related to posterior septum of the knee joint and arthroscopic surgery was extensively reviewed and analyzed. Results The posterior septum of the knee joint has more mechanoreceptors and blood vessels in the upper part, which are close to arteria popliteal at the tibial plateau level; the posterior compartment is divided into wider posteromedial and narrower posterolateral compartments. A safe arthroscopic trans-septal portal is established, in the knee flexion of 90°, in a lateral-to-medial direction, and with an inserting location below the middle of posterior septum. Conclusion The establishment method of posterior trans-septal portal is not uniform and all the features of posterior septum should be considered to decrease the complications.