Changes in glycosylation can serve as markers for rare genetic disorders, including lysosomal storage diseases (LSDs). Nephropathic Cystinosis (NC), caused by mutations in the CTNS gene, is characterised by cystine accumulation in lysosomes due to dysfunctional cystinosin, a heavily N-glycosylated lysosomal transporter. We analysed total serum and IgG N-glycosylation using hydrophilic interaction ultra performance liquid chromatography (HILIC-UPLC) to explore the diagnostic biomarker capabilities and their pathophysiological relevance in NC. In this double-blind study (n = 12), we examined N-glycosylation of total serum and serum IgG from Irish participants with and without NC. Dimensionality reduction methods were used applying their glycan data to predict NC status, yet only modest predictive power was observed (66.6% for serum and 50% for IgG N-glycosylation). However, upon unblinding the data, we identified significant differences in specific serum N-glycosylation in NC, particularly in sialylation. These findings provide the first evidence that serum N-glycosylation is altered in NC. These changes may indicate disease-associated systemic alteration including dysregulation in N-glycosylation pathway. It provides justification for the need for a larger validation study and invites further exploration of its role in NC pathophysiology. We provide key recommendations for age stratification for studying serum, plasma and IgG N-glycans in juvenile cohorts as they display unique profiles compared to adult populations, an important consideration for all juvenile studies, even beyond the scope of rare diseases.