No previous studies reported the association of IL-12 polymorphisms with osteosarcoma. We aimed to investigate the association in a Chinese population. IL-12A rs568408, rs2243115, and IL-12B rs3212227 polymorphisms were evaluated in a case-control study of 106 osteosarcoma patients and 210 health controls by using polymerase chain reaction-restriction fragment length polymorphism. Serum IL-12p40 levels were measured by enzyme-linked immunosorbent assay. The serum IL-12p40 levels were significantly higher in controls than those in osteosarcoma patients (p<0.01). Genotypes of rs568408 GA and GA/AA, and rs3212227 CC and AC/CC were associated with the risk of osteosarcoma (rs568408 GA: odds ratios [OR]=1.86, 95% confidence intervals [CI]=1.11-3.12; GA/AA: OR=1.75, 95% CI=1.06-2.89, and rs3212227 CC: OR=2.70, 95% CI=1.38-5.28; CC/AC: OR=1.73, 95% CI=1.03-2.90). Moreover, rs3212227 CC/AC genotypes were significantly associated with decreased serum IL-12p40 levels in osteosarcoma patients compared to AA genotypes (p=0.035). Stratification analysis showed no associations between rs3212227 variant and the patients’ gender, tumor location, and metastasis. Our data suggest that the serum IL-12p40 levels associate with the risk of osteosarcoma and are regulated by IL-12B rs3212227 polymorphism. The IL-12A rs568408 and IL-12B rs3212227 may confer the susceptibility to osteosarcoma risk.