The objectives of this pilot-scale study were to (1) evaluate the effectiveness of bioremediation of trichloroethylene (TCE)-contaminated groundwater with the supplement of slow polycolloid-releasing substrate (SPRS) (contained vegetable oil, cane molasses, surfactants) under reductive dechlorinating conditions, (2) apply gene analyses to confirm the existence of TCE-dechlorinating genes, and (3) apply the real-time polymerase chain reaction (PCR) to evaluate the variations in TCE-dechlorinating bacteria (Dehalococcoides spp.). Approximately 350 L of SPRS solution was supplied into an injection well (IW) and groundwater samples were collected and analyzed from IW and monitor wells periodically. Results show that the SPAS caused a rapid increase of the total organic carbon concentration (up to 5794 mg/L), and reductive dechlorination of TCE was significantly enhanced. TCE dechlorination byproducts were observed and up to 99% of TCE removal (initial TCE concentration = 1872 mu g/L) was observed after 50 days of operation. The population of Dehalococcoides spp. increased from 4.6 x 10(1) to 3.41 x 10(7) cells/L after 20 days of operation. DNA sequencing results show that there were 31 bacterial species verified, which might be related to TCE biodegradation. Results demonstrate that the microbial analysis and real-time PCR are useful tools to evaluate the effectiveness of TCE reductive dechlorination. (C) 2014 Elsevier B.V. All rights reserved.