An anammox attached film expanded bed (AAFEB) reactor was operated to study the long-term performance and the variation of substrate tolerance ability. The results indicated that the nitrogen loading potential (NLP) was significantly enhanced from 13.56 gN.(L.d)(-1) to 20.95 gN.(L.d)(-1) during the stable operation period. The inhibitory concentration of 10% (IC10) for free ammonia (FA), free nitrous acid (FNA) and SNinf (diluted substrate concentration) increased from 18 mg/L, 12 mu g L-1 and 370 mg NL-1 to 31 mg/L, 19 mu g L-1 and 670 mg NL-1, respectively. However, the substrate shock of 2500 mg NL-1 for 24 h terribly weakened the treatment performance and substrate tolerance ability of the reactor. The results of batch tests indicated that the existence of lag phase made the AAFEB reactor more vulnerable to substrate variation. The SNinf was accurate to be used to monitor the reactor performance and should be maintained below 320 mg NL-1 to ensure the absolute stable operation. (C) 2016 Elsevier Ltd. All rights reserved.