In order to enhance the use of biofilm models for simulation of wastewater treatment plants (WWTPs), model calibration is absolutely necessary. The aim of this work was to develop a calibration protocol for biofilm processes model with a help of the sensitivity analysis of the most important parameters implemented in BioWin® software. A case study of a circulating fluidized bed bioreactor (CFBBR) system used for biological nutrient removal (BNR) from municipal wastewater was used to verify and validate the proposed calibration protocol. Applying the various stages of the proposed biofilm calibration procedures enhanced the applicability of BioWin®, which was capable of predicting most of the performance parameters with an average percentage error (APE) of 0% to 20%. Although the calibrated model predicted most of the performance parameters, BioWin® predicted the total biomass and SRT based on active biomass only, whereas in the CFBBR both active as well as inactive biomass accumulated.