Introduction: Advanced stage neuroblastoma is associated with 60-70% mortality. We propose that expression microarrays can be used to effectively identify novel biomarkers and targets for high risk disease.Methods: Using a cDNA microarray comparing tumor tissue from high risk versus low risk disease, an expression sequence tag (EST) over-expressed in high risk neuroblastoma was identified and found to correlate with a candidate gene designated as neuroblastoma secreted protein (NDSP). Northern blot, reverse transcriptase polymerase chain reaction (RT-PCR), and quantitative PCR were used to screen expression in cell lines and tissue samples. NDSP was cloned into an expression vector with a C-terminus V5-tag (NDSP-V5) and transfected into HEK293T cells for Western analysis. A polyclonal antibody (anti-NDSP-Ab) was generated to test endogenous protein synthesis in neuroblastoma.Results: The NDSP gene is located on chromosome 1q24.3 and consists of a 501 base pair open reading frame encoding a 167 amino acid peptide. NDSP is expressed in neuroblastoma, retinoblastoma, and neuroepithelioma; however, expression is not found in normal human tissues or 10 other cancer types tested. Additionally, NDSP is expressed in 9 neuroblastoma tissue samples representing stages 1-4 and 4S. Quantitative PCR revealed that NDSP over-expression correlates with high risk disease (stage 3 and 4). Western blot of cell lysate and supernatant from NDSP-V5 transfected cells shows a 20 kDa band. Additional Western blots with anti-NDSP-Ab showed abundant NDSP protein levels within all neuroblastoma cell lines tested.Conclusions: NDSP is a novel protein biomarker identified by microarray analysis and over-expressed in high risk neuroblastoma.