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The mechanical properties of micro‐ and nanoscale materials directly determine the reliability of heterostructures, microstructures, and microdevices. Therefore, an accurate evaluation of the 3D strain field at the nanoscale is important. In this study, a scanning transmission electron microscopy (STEM) moiré depth sectioning method is proposed. By optimizing the scanning parameters of electron probes at different depths of the material, the sequence STEM moiré fringes (STEM‐MFs) with a large field of view, which can be hundreds of nanometers obtained. Then, the 3D STEM moiré information constructed. To some extent, multi‐scale 3D strain field measurements from nanometer to the submicrometer scale actualized. The 3D strain field near the heterostructure interface and single dislocation accurately measured by the developed method. [ABSTRACT FROM AUTHOR]