Chondroitin sulfate (CS) and its isomeric variant, dermatan sulfate (DS), are complex glycosaminoglycans (GAGs) which are ubiquitous components of the extracellular matrix in various tissues including the brain. CS and/or DS are known to bind to a variety of growth factors and regulate many cellular events such as proliferation and differentiation. Although the biological activities of CS and/or DS towards neural stem/progenitor cells (NSPCs) have been well investigated, the CS and/or DS of hematopoietic stem cells (HSCs) have not been fully characterized. Here, we analyzed GAGs on mononuclear cells of rat umbilical cord blood cells (UCB-MNCs). CS was detected in vascular intima and media of rat umbilical cord at embryonic day 19 (E19) by immunohistochemistry. The stem-cell-enriched-UCBCs (SCE-UCBCs), which were expanded from rat UCB-MNCs, expressed CS. CS chains are composed of repeating disaccharide units, which are classified into several types such as O-, A-, B-, C-, D-, and E-unit according to the number and positions of sulfation. A disaccharide composition analysis revealed that CS and/or DS were abundant in rat UCB-MNCs as well as in their expanded SCE-UCBCs, while the amount of heparan sulfate (HS) was less. The degree of sulfation of CS/DS was relatively low and the major component in UCB-MNCs and SCE-UCBCs was the A-unit. A colony-forming cell assay revealed that the percentage of colony-forming cells decreased in culture with CS degradation enzyme. The CS and/or DS of UCBCs may be involved in biological activities such as stem cell proliferation and/or differentiation.