Aim: Ankylosing spondylitis (AS) is a chronic systemic inflammatory rheumatic disorder that primarily affects the axial skeleton. To date there is no satisfactory biomarker for diagnosis or prognosis of AS. This study is focused on detection of potential biomarkers for the diagnosis of AS using surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF-MS) with the ultimate goal of developing a progression biomarker for AS. Materials & methods: Protein profiles of sera of 38 AS patients and 38 healthy controls were obtained using ProteinChip arrays. Different arrays and conditions were compared with find out which condition resulted in the optimal discrimination of both groups. Results: Optimal discrimination was reached using weak cation exchange (CM10) and immobilized metal affinity capture coupled with copper (IMAC-Cu2+) arrays. Using a multimarker approach on protein profiles obtained with CM10 arrays resulted in a sensitivity of 66% and a specificity of 74%. Using a multimarker approach on protein profiles obtained with IMAC-Cu2+ arrays resulted in a sensitivity and specificity of 70%. Conclusions: This is the first study that shows that protein profiling in serum using SELDI-TOF-MS can be used as a diagnostic tool for AS. The biomarkers will be validated to confirm if they can be used as progression biomarkers.