This study aims to detect S100A12 in serum and synovial fluid (SF) of knee osteoarthritis (OA) patients and to investigate their correlation with clinical severity of the disease. This study enrolled 198 OA patients and 84 healthy controls. The radiological/symptomatic severity of the disease was assessed by the Kellgren-Lawrence (KL) grading system/Western Ontario McMaster University Osteoarthritis (WOMAC) scores. We demonstrated that S100A12 levels in SF but not serum were independently associated with KL grades and WOMAC scores. These results suggested that the S100A12 in SF might be a potential biomarker for predicting the development and progression of knee OA.