ISOBAM-104 stabilized Fe colloidal catalysts were synthesized by in-situ reduction method for hydrogen generation from KBH4 hydrolysis in alkaline solutions. The as-prepared non-noble metal colloidal catalysts contained Fe nanoparticles with an average size of around 1.9 nm as well as uniformly dispersed single Fe atoms, exhibiting a high catalytic activity of 8100 mL-H2 min-1 g-Fe- 1, which was higher than that of the Fe catalysts prepared by other protective agents. Such high catalytic activity of the as-synthesized Fe colloidal catalysts could be ascribed to the following two factors: (1) the in-situ formed and well-dispersed Fe atoms that were confirmed by HAADFSTEM, and (2) the superior protective ability of ISOBAM-104 compared with other protective agents, which was validated by a density functional theory (DFT). In addition, the apparent activation energy of Fe colloidal catalysts was 49.85 kJ mol-1, which was in coincidence with DFT calculation results (49.7 kJ mol-1) and lower than most other Fe-based catalysts, implying that the as-prepared Fe colloidal catalyst is a promising candidate for the catalytic hydrolysis of KBH4. This study could provide a promising approach to prepare non-noble metal colloidal catalysts with excellent catalytic activity for hydrogen generation.