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Manganese oxide is an effective promoter for enhancing the activity and selectivity of cobalt-based Fischer-Tropsch synthesis (FTS) catalysts. Previous studies suggest that Co atoms located at the Co-MnO perimeter adsorb CO more strongly and undergo C-O bond cleavage more readily than CO adsorbed on Co sites, which leads to improved C5+ selectivity. Thus, direct characterization of the formation and structure of the active interface between Co nanoparticles and MnO is essential for a full understanding of the FTS promotion effect. Scanning transmission electron microscopic (STEM) methods are well-suited for direct characterization of nanoscale features through imaging, energy dispersive spectroscopy (EDS) elemental mapping, and spatially resolved electron diffraction, a technique known as scanning nanobeam diffraction. However, these techniques require that the catalytic nanoparticles are non-overlapping and well-defined, which is not typical for nanoparticles on porous metal oxide FTS supports. In the present study, we addressed these challenges by synthesizing catalysts composed of cobalt dispersed on nonporous alumina nanospheres (Co/ANS), which yielded isolated nanoparticles that are amenable to elemental and phase mapping by STEM-EDS and scanning nanobeam diffraction. Then, manganese oxide-promoted Co/ANS (Mn-Co/ANS) was characterized using these two techniques, which provided direct evidence for the transformation of a Co-Mn oxide spinel present in the as-prepared catalyst into closely associated Co3O4 and MnO2 nanoparticles in the passivated catalyst after its use for FTS.