The search for new biomarkers for neuroblastoma diagnosis is driven by the need for improved accuracy, early detection, and personalized treatment of this particular type of cancer. Neuroblastoma is a pediatric cancer that primarily affects young children, and its diagnosis can be challenging due to its diverse clinical presentation and variable outcomes. Small extracellular vesicles (sEVs) have gained attention as potential tumor biomarkers due to their unique characteristics and their ability to reflect the molecular profile of the parent tumor cells. sEVs released by tumor cells can carry a cargo of proteins, nucleic acids (such as DNA and RNA), and other molecules that reflect the molecular diversity of the tumor. By analyzing the content of sEVs, researchers can gain insights into the heterogeneity of the tumor and potentially identify specific biomarkers associated with tumor subtypes or disease progression. sEVs can be found in various body fluids, this accessibility makes sEVs a promising source for non-invasive or minimally invasive diagnostic and monitoring approaches. sEVs are protected by a lipid bilayer membrane, which shields their cargo from degradation by enzymes and other harsh conditions in the extracellular environment. This stability allows sEVs to circulate in body fluids for extended periods, making them suitable for biomarker analysis. Here, we performed mass spectrometry-based proteomic analysis of serum sEVs to identify and validate potential protein biomarkers for diagnosis of NB.