Electrophoretic analysis of N-glycans on microfluidic devices.
We report rapid and efficient electrophoretic separations of N-glycans on microfluidic devices. Using a separation length of 22 cm and an electric field strength of 750 V/cm, analysis times were less than 3 min, and separation efficiencies were between 400,000 and 655,000 plates for the N-glycans and up to 960,000 plates for other sample components. These high efficiencies were necessary to separate N-glycan positional isomers derived from ribonuclease B and linkage isomers from asialofetuin. Structural isomers of N-glycans derived from a blood serum sample of a cancer patient were also analyzed to demonstrate that clinically relevant, complex samples could be separated on-chip with efficiencies similar to those derived from model glycoproteins. In addition, we compared microchip and capillary electrophoresis under similar separation conditions, and the microchips performed as well as the capillaries. These results confirmed that the noncircular cross section of the microchannel did not hamper separation performance. For all experiments, the glycan samples were derivatized with 8-aminopyrene-1,3,6-trisulfonic acid to impart needed charge for electrophoresis and a fluorescent label for detection.
Department of Chemistry and the National Center for Glycomics and Glycoproteomics, Indiana University, Bloomington, Indiana 47405-7102, USA.
Zexi
Z
Zhuang
Jason A
JA
Starkey
Yehia
Y
Mechref
Milos V
MV
Novotny
Stephen C
SC
Jacobson
D001212:Asialoglycoproteins / Q000737:chemistry; D004586:Electrophoresis; D060748:Fetuins; D007536:Isomerism; D044085:Microfluidics / Q000295:instrumentation* / Q000379:methods*; D008956:Models, Chemical; D011134:Polysaccharides / Q000032:analysis* / Q000737:chemistry*; D012260:Ribonucleases / Q000737:chemistry; D000509:alpha-Fetoproteins / Q000737:chemistry
D001212:Asialoglycoproteins; D060748:Fetuins; D011134:Polysaccharides; D000509:alpha-Fetoproteins; C017029:asialofetuin; D012260:Ribonucleases; C029600:ribonuclease B