This study investigates the mechanisms of temper embrittlement in 410 martensitic stainless steel, a material widely used in steam turbine blades due to its excellent corrosion resistance and high strength achieved through quenching and tempering heat treatments. While the material’s hardness and impact toughness strongly depend on tempering temperatures, significant embrittlement occurs around 540°C, manifesting as decreased Charpy impact energy alongside increased strength and hardness. To understand this phenomenon at the nanometer scale, high-resolution transmission electron microscopy (TEM) analysis was performed, focusing on electron diffraction patterns along the <110>α-Fe and <113>α-Fe zone axes. The analysis revealed distinctive double electron diffraction spots at 1/3(211) and 2/3(211) positions, with lattice spacing of approximately 3.5 Å—triple the typical α-bcc lattice spacing (1.17 Å). These regions were identified as metastable “zones” resembling ω-phase structures, potentially responsible for the embrittlement. While this newly identified phase structure may not fully explain the complex mechanisms of temper embrittlement, it provides valuable insights for developing improved alloying and heat treatment methods to mitigate embrittlement in martensitic steels.