Structural stability is highly important for the safe evacuation of occupants and safe access to fire-fighters in accidental fire events. In this paper, the effect of fire protection of beams on the fire resistance of heated columns is studied firstly, and the results show that fire protection of beams may have little effect on the behaviours of heated columns as the heating time of fire spread scenario is short in this study. Then, the behaviours of 3D steel frames subjected to seven different fire spread scenarios have been investigated. The effects of fire source location, occurrence time of fire spread, and direction of fire spread on the fire resistance of steel frame are comprehensively studied. In case of horizontal fire spread scenarios with different fire source compartments, collapse occurs when the corner and long edge compartments are fire source compartments. A stable catenary mechanism of beam could improve and control the collapse time and range. The corner column may bear the catenary tensions in two horizontal directions, and the lateral stiffness of corner column should be crucial for the design of structural fire. For the non-insulated steel frame, the maximum temperature and maximum axial force of heated columns appear at same time and collapse finally happens in the scenario that fire spread when the gas temperature of fire source compartment reaches the peak temperature. The time of most adverse spread for the non-insulated steel frame should be that when the gas temperature of fire source compartment reaches the peak temperature. The influence of vertical fire spread on heated columns may be smaller than that of horizontal fire spread, but vertical fire spread had great influence on fire resistance of beam. According to this study, spread range, duration, and fire temperature are the three most critical factors of fire spread, and the adverse horizontal spread scenario could be mainly selected from these factors.