Spatially resolved analysis of magnetic properties on the nanoscale remains challenging, yet strain and defects on this length-scale can profoundly affect a material’s bulk performance. We present a detailed investigation of the magnetic properties of La0.67Sr0.33MnO3 thin films in both free-standing and nanowire form and assess the role of strain and local defects in modifying the films’ magnetic properties. Lorentz transmission electron microscopy is used to measure the magnetocrystalline anisotropy and to map the Curie temperature and saturation magnetization with nanometric spatial resolution. Atomic-scale defects are identified as pinning sites for magnetic domain wall propagation. Measurement of domain wall widths and crystalline strain are used to identify a strong magnetoelastic contribution to the magnetic anisotropy. Together, these results provide unique insight into the relationship between the nanostructure and magnetic functionality of a ferromagnetic complex oxide film.