A surgical access port for passage through body tissue to provide access to an underlying tissue site is provided. The access port has a working channel dimensioned for receiving a surgical instrument and a seal assembly for providing a substantial seal before, during and after insertion of a surgical instrument. The seal assembly includes an instrument seal and a zero seal. The instrument seal includes a proximal base that is interconnected to a distal instrument engaging portion by an elongate supporting portion. The engaging portion has a lateral dimension larger than the lateral dimension of the supporting portion forming a bulbous, mushroom-like head. The lateral dimension of the engaging portion decreases toward a distal opening of the instrument seal such that the distal end of the engaging portion is substantially perpendicular to the seal axis. The supporting portion closely conforms to an inserted instrument and serves to align the distal end of the seal.