The use of supporting media for the immobilization of microorganisms is widely known to provide a surface for microbial growth and a shelter that protects the microorganisms from inhibitory compounds. In our previous studies, activated carbon (AC) alone used as a support medium for H2S biological removal was proved prompt and efficient in a bench-scale biofilter and biotrickling filter. In this study, the mechanisms of H2S elimination using microbial immobilized activated carbon, i.e., biological activated carbon (BAC), are investigated. A series of BAC as supporting medium were taken from the inlet to outlet of a bench-scale horizontal biotrickling filter to examine the different effects of physical/chemical adsorption and microbial degradation on the overall removal of H2S. The surface properties of BAC together with virgin and exhausted carbon (after H2S breakthrough test, non-microbial immobilization) were characterized using the sorption of nitrogen (Braunner-Emmett-Teller test), scanning electron microscopy (SEM), surface pH, thermal, carbon-hydrogen-nitrogen-sulfur (CHNS) elemental and Fourier transform infrared (FTIR) analyses. Tests of porosity and surface area provide detailed information about the pore structure of BAC along the bed facilitating the understanding of potential pore blockages due to biofilm coating. A correlation between the available surface area and pore volume with the extent of microbial immobilization and H2S uptake is evidenced. SEM photographs show the direct carbon structure and biofilm coated on carbon sur-FTIR spectra, differential thermogravimetric curves and CHNS results indicate less diversity of H2S oxidation products on BAC than those previously observed on exhausted carbon from H2S adsorption only. The predominant oxidation product on BAC is sulfuric acid, and biofilm is believed to enhance the oxidation of H2S on carbon surface. The combination of biodegradation and physical adsorption of using BAC in removal of H2S could lead to a long-term (i.e., years) good performance of biotrickling filters and biofilters based on BAC compared to carbon adsorption only.