Laser powder bed fusion (PBF) has been used to create structures of many different alloys including stainless steels (SS). In contrast to SS prepared via conventional approaches, PBF-produced SS often contains a high concentration of oxygen. It is generally believed that the oxygen primarily exists as oxide inclusions with diameters ranging from tens of nanometers to several microns. Thermodynamic calculations also show that the solubility of the oxygen is extremely low in the liquid, face center cubic (FCC), or body center cubic phases that are relevant to the composition of the SS investigated in this study. Additionally, these calculations predict that majority of oxygen stays in the metastable MnSiO₃ phase. In this study, we perform multi-scale, quantitative electron microscopic analysis on the as-printed SS 316L and find that a large amount of oxygen actually exists in the interstitial sites of the alloy lattice, suggesting that oxygen might have been trapped in the alloy substrate during the rapid cooling process of PBF. The observations from the atomic-scale-resolution characterization are supported by the first principles simulations through density functional theory calculations, which reveal that oxygen can stay energetically stable in the octahedral site of the FCC structure. Additionally, these interstitial oxygen atoms can form ionic bonds with the neighboring metal atoms, with a particularly high affinity towards Cr atoms. The presence of interstitial oxygen in the SS substrate appears to assist the surface passivation processes and lead to the exceptionally high pitting potential. The identification of a large amount of interstitial oxygen in the alloy may have a profound impact on the design of strong, tough, and corrosion resistant alloys.