This prospective, randomized, controlled clinical trial assessed the therapeutic effects of major ozone autohemotherapy (O3-MAH) in patients with post-acute sequelae of COVID-19 (PASC). Seventy-three eligible participants were randomly assigned to an O3-MAH plus conventional therapy group (n = 35) or a conventional therapy alone group (n = 38). Symptom score, pulmonary function, 6-minute walk distance (6MWD), and hematological, biochemical, and immunological parameters were evaluated before and after the interventions. Both groups demonstrated improvements in various parameters post-intervention, but efficacy was greater in the O3-MAH group than the conventional treatment group; with intervention effectiveness defined as a >= 50 % reduction in symptom score, 25 of 35 patients (71 %) responded to O3-MAH, while 17/38 patients (45 %) responded to conventional treatment alone (P = 0.0325). Significant improvements in symptom scores (P = 0.0478), tidal volume (P = 0.0374), predicted 6MWD (P = 0.0032), and coagulation and inflammatory indicators were noted in the O3-MAH group compared with the conventional treatment group. O3-MAH was more likely to be effective in patients with elevated CRP levels. Furthermore, O3-MAH markedly improved cellular immunity, and this improvement became more pronounced with extended treatment duration. In summary, combining O3-MAH with conventional treatment was more effective than conventional therapy alone in improving symptoms, pulmonary function, inflammation, coagulation, and cellular immunity in patients with PASC. Further research is now warranted to validate these findings and individualize the regimen.