Several strategies with different combination of inocula and packing material were investigated to obtain the opti-mal start-up time and elimination capacity (EC) in toluene biofiltration. The inocula contained the activated sludge and tolu-ene degrading bacteria, and the packing material consisted of different mixing ratios of peat and wood chips. A final toluene load of 21.2 g/(m3·h) was attained step by step in four parallel biofilters. A shortest start-up time of 15 days and a highest EC of 17.0 g/(m3·h) were observed in the biofilter B-4, which was inoculated with a special microbial consortium consisting of three strains of toluene degrading bacteria and was packed with the mixture of peat and wood chips at a ratio of 80:20 (w/w). These results indicated that inoculating pre-acclimatized microbes dramatically shortened the start-up time, and such a com-position of packing material could maintain an appropriate environment (with the bed porosity and water content equating to 0.45 and 1.96, respectively) for the growth of dominant toluene degrading bacteria in the biofilter.