The purpose of this paper is to refine the BIOMATH calibration protocol for SBR systems, in particular to develop a pragmatic calibration protocol that takes advantage of SBR information-rich data, defines a simulation strategy to obtain proper initial conditions for model calibration and provides statistical evaluation of the calibration outcome. The updated calibration protocol is then evaluated on a case study to obtain a thoroughly validated model for testing the flexibility of an N-removing SBR to adapt the operating conditions to the changing influent wastewater load. The performance of reference operation using fixed phase length and dissolved oxygen set points and two real-time control strategies is compared to find optimal operation under dynamic conditions. The results show that a validated model of high quality is obtained using the updated protocol and that the optimization of the system's performance can be achieved in different manners by implementing the proposed control strategies.