Tunable diode laser absorption spectroscopy has proven to be highly advantageous in the diagnostics of high enthalpy air plasma due to its exceptional sensitivity, selectivity, rapid response, and adaptability to harsh environments. However, measuring dissociated atoms remains challenging because these atoms are short-lived reactive species that quickly react with surrounding particles or collide to form stable molecules, making it difficult to prepare an atomic gas source capable of generating sustained, concentration-stabilized, and high signal-to-noise absorption spectroscopy. In this study, we have developed a laboratory-based glow discharge plasma static calibration system designed for time-resolved measurements of the number density of specific electronic states of O atoms. Experimental results indicate that a low-pressure glow discharge of NO can generate a substantial number of metastable O atoms at relatively stable concentrations. The concentration of these metastable O atoms within the discharge tube can be controlled by adjusting gas pressure and discharge current. As the gas pressure in the discharge tube increases, the concentration of metastable O atoms rises to a peak value of 3.66 × 109 cm−3 before declining. The system’s stability was assessed using Allan variance analysis, revealing the detection limit of metastable O atom number density is 8.0018 × 106 cm−3 when the average time is 3.2 s. By varying the input gas, the system is also capable of generating significant quantities of stable N and OH radicals. The system’s stability, controllability, and versatility in producing reference gases of known composition make it a reliable tool for diagnosing high enthalpy flow fields.