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The high residual stress (RS) in the components manufactured by laser powder bed fusion (LPBF) can be detrimental to the mechanical properties, especially the fatigue performance. Stress relief (SR) heat treatment is commonly applied on the as-built components for stress relaxation. Hence, the effect of the SR heat treatment (600 C/4 hours) on the fully reversed low cycle fatigue (LCF) behaviours of LPBF stainless steel 316L (SS 316L) was investigated in this study. The depth profiles of RS distribution across the specimen were measured by neutron diffraction, and a significant RS relaxation (nearly 46%) was obtained after the SR heat treatment. The compressive RS at the centre of the specimen induces an asymmetrical responding stress. Nearly identical microstructure and dislocation structure were observed between the as-built (AB) and the SR specimens, along with a similar fatigue strain-life. However, a higher responding stress (by approximately 50 MPa) is found in the SR specimens, which indicates a better fatigue stress-life. The higher responding stress of the SR specimen results from the better cyclic hardening capability and the higher strength in the elasto-plastic regime. The commonly observed cell structure in LPBF SS 316L evolves toward a wall-like dislocation structure via planar dislocation movement due to the plasticity induced during cyclic loading. At the lower applied strain ranges, the strain induced α’martensitic transformation was observed at the cell boundaries, which is an inverse trend compared to the wrought SS 316L. Yet, the α’martensite does not show significant contribution to the mechanical properties.