Sialic acids often terminate glycan structures and are believed to have significant functions in cancer development and metastasis. Understanding how sialylation is affected by disease onset and progression will provide critical information for clinical diagnosis and prognosis. To study sialylation patterns in clinical samples, we neutralize the negative charge on a2,3- and a2,6-linked sialic acids by covalent modification. After neutralization and fluorescent labeling with 8-aminopyrene-1,3,6-trisulfonic acid, all glycans in the samples have the same effective charge and, consequently, their separation is based on hydrodynamic volume. Glycan structures are resolved by microchip electrophoresis on a 22-cm separation channel operated at 1250 V/cm and detected by laser-induced fluorescence. With our neutralization strategy, we are able to identify which glycans in complex mixtures are sialylated and directly compare electrophoretic mobilities from these separations to the mass-to-charge ratios obtained from MALDI-MS analysis of the same samples. Moreover, we conserve the information provided by the sialic acids and are able to identify sialylated and non-sialylated structures without enzymatic sequencing. We can also determine if highly charged, low molecular weight glycans, which are difficult to characterize because their mobilities can be similar to structures associated with the fluorescent labeling reaction, are present in serum samples. If present, we will be able to effectively monitor these glycans and discern their roles in disease.

Analysis of sialylated glycans by microchip electrophoresis

246th National Meeting (2013)

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