This study explores the relationship between the microstructure, composition, orientation, and mechanical properties of the nickel-based superalloy Inconel 718. Employing scanning electron microscopy (SEM), electron microprobe (EMP), nano-indentation, and other techniques, the study observes the structure, confirms the composition, determines orientation, and tests mechanical properties in specific micro-zones. Findings reveal a uniform grain distribution in Inconel 718, with a minor δ-phase presence at grain boundaries. There is a notable enrichment of Nb at the grain boundaries, whereas Fe and Cr levels are lower at these boundaries compared to the grain interiors. The indentation hardness and modulus at the grain boundaries are markedly higher than those within the grains. Moreover, grains with different orientations exhibit diverse microscale mechanical properties, such as hardness and elastic modulus. This research establishes a quantitative mapping relationship between the microstructure, composition, orientation, and mechanical properties of Inconel 718, providing a foundation for future multiscale (micro to macro) mechanical property investigations.