Osteomyelitis (OM) is a major challenge in orthopedic surgery. The diagnosis of OM is based on imaging and laboratory tests, but it still presents some limitations. Therefore, a deeper comprehension of the pathogenetic mechanisms could enhance diagnostic and treatment approaches. OM pathogenesis is based on an inflammatory response to pathogen infection, leading to bone loss. The present study aims to investigate the potential diagnostic role of a panel of osteoimmunological serum biomarkers in the clinical approach to OM. The focus is on the emerging infection biomarker sCD14-ST, along with osteoimmunological and inflammatory serum biomarkers, to define a comprehensive biomarker panel for a multifaced approach to OM. The results, to our knowledge, demonstrate for the first time the diagnostic and early prognostic role of sCD14-ST in OM patients, suggesting that this biomarker could address the limitations of current laboratory tests, such as traditional inflammatory markers, in diagnosing OM. In addition, the study highlights a relevant diagnostic role of SuPAR, the chemokine CCL2, the anti-inflammatory cytokine IL-10, the Wnt inhibitors DKK-1 and Sclerostin, and the RANKL/OPG ratio. Moreover, CCL2 and SuPAR also exhibited early prognostic value.