In order to prove that perchlorate-laden resins could be bio-regenerated through direct contact with perchlorate-reducing bacteria (PRB), a combined bio-regeneration and ion-exchange (IX) system was operated. Two kinds of perchlorate-laden resins, nitrate-selective A520E and perchlorate-selective A530E, were successfully regenerated by PRB cultivated under anaerobic conditions. The bio-regeneration efficiency of perchlorate-laden resins increased with the amount of flow passed through the IX column. When the fully exhausted resin was bio-regenerated for 10 days at the flow rate of 2 BV (bed volume)/min and mixed liquor suspended solids concentration of 80 mg/L, almost 100% of IX capacity was recovered. A520E resin had higher bio-regeneration efficiency than A530E under all conditions, probably due to the fact that the perchlorate ion is more strongly bonded to the functional group of perchlorate-selective A530E resin. Measurement of perchlorate concentrations in the column effluents also revealed that the amount of perchlorate eluted from A520E resin was higher than that from A530E resin. Since only 10–20% of perchlorate was eluted from the resin during 10 days of bio-regeneration, the main mechanism of bio-regeneration appears to be the direct reduction of perchlorate by PRB on the resin.