A multi-step response surface methodology was successfully applied to interpret the adsorption characteristics of lead, cadmium and copper ions onto immobilized spent Tricholoma lobayense (ISTL) in single and ternary systems in this study. Firstly, the most significant medium factors, which were pH, biomass loading and contact time, on biosorption of three metals, were determined by Minimum Run Res V Design. Then central composite design followed by mixture design was utilized to identify the affinity and predict the adsorption capacities of three metals onto the biosorbent. The results showed that the preference of ISTL for three metals was in the order of Pb(II) > Cu(II) > Cd(II) and lead ions could still be effectively removed from aqueous solution in the presence of both cadmium and copper while removal of the cadmium and copper ions would be suppressed by lead. Sorption isotherm data were better represented by Freundlich model. The both adsorption and desorption process of three metals followed the pseudo-second-order kinetics model. Scanning electron microscope (SEM) and energy dispersive spectrometry (EDS) analysis testified the obvious change of the surface morphology and the presence of three metals on the biosorbent after metal binding. The ISTL could be regenerated with 1 M HNO3, which allowed the reuse of the biomass in three biosorption-desorption cycles without considerable loss of adsorption capacity. The present work suggests that ISTL as an abundant low-cost biomaterial was an efficient biosorbent for heavy metals removal from wastewater. (C) 2010 Elsevier B.V. All rights reserved.