Studies on anaerobic ammonium oxidation (ANAMMOX) have been increasing due to its advantages over traditional nitrification–denitrification systems. However, long and difficult start-up periods of ANAMMOX reactors may sometimes be frustrating for researchers dealing with them first time in the lab. Therefore, a guide for starting up ANAMMOX reactors will greatly help them and enhance the use of this technology. Three trials, each has three reactors, have been carried out to reach successful start up in this study. An expected ANAMMOX enrichment could not be achieved in the first and second trials. Under the light of the experiences gained from the unsuccessful ones, the third trial resulted in >95% ammonium removals within 20–50 days. It is possible to startup or to enrich ANAMMOX reactors within 50 days by presented strategy guide.