Labeling conditions using a 2-aminobenzamide reagent for quantitative analysis of sialo-oligosaccharides

During 2-aminobenzamide (2-AB)-labeling of sialo-oligosaccharides the autho rs encountered a significant loss of sialic acids under the reported condit ions, at 65 degrees C for 2 h. By examination of the relationship between l abeling temperature and recovery of tetra-sialylated oligosaccharides, 2-AB -labeling at 37 degrees C far 16 h resulted in a maximum yield with practic ally no loss of sialic acid. Under the newly optimized conditions, the reproducibility of 2-AB-labeling was examined in triplicate with three lots of recombinant human erythropoie tin (r-HuEPO). The present conditions gave analytical data comparable to th ose obtained by the established NaB[H-3](4) labeling procedure. The modifie d method has been applied to oligosaccharide structural analysis of sialogl ycoprotein therapeutics.