This study examined the effect of playing matches against different opponent teams (hard match; HM, medium match; MM, and easy match; EM) on pre-match testosterone concentration (T), pre-to-post match cortisol (C) concentration, and pre-competitive anxiety (CSAI-2 questionnaire) in 20 elite male basketball players. One training session (TS) was also assessed (control condition). Saliva steroids were determined by ELISA. The pre-T concentration was significantly lower for the TS (163±54pmolL), compared to the three official matches (EM: 208±82pmolL, MM: 213±57pmolL, and HM: 218±37pmolL) (p<0.05). A significant change in C from pre-to-post was observed for all conditions (TS [5.8±3.7 vs 14.4±9.4ƞmolL], EM [10.1±4.9 vs 17.3±10.2ƞmolL], MM [13.0±7.4 vs 21.5±8.3ƞmolL], and HM [18.9±5.8 vs 31.4±5.3ƞmolL] (p<0.05). A higher PRE-C was observed for the HM, compared to TS and EM (p<0.05) and a higher POST-C concentration for the HM was detected, compared to the TS, EM, and MM (p<0.05). A lower anxiety level was observed for the EM (somatic: 15.0±3.7; cognitive: 15.8±4.5 arbitrary units [AU]) compared, to the MM (somatic: 16.4±3.5; cognitive: 17.7±4.0AU) and HM (somatic: 15.8±3.9; cognitive: 18.3±3.9AU) (p<0.05) and a higher self-confidence was observed for the TS (28.9±5.1AU), compared to the HM (26.7±3.0AU) (p<0.05). These results suggest that playing against a high-level opponent may cause a higher psychobiological stress, likely because opponent level may be perceived as threating to the social status in a given hierarchy.