Hyphenated technique for releasing and MALDI MS analysis of O-glycans in mucin-type glycoprotein samples

Anal Chem. 2010 Sep 1;82(17):7436-43. doi: 10.1021/ac101581n.

Abstract

We developed an automatic apparatus for the release of O-glycans from mucin-type glycoproteins and proteoglycans (Matsuno, Y.-k.; Yamada, K.; Tanabe, A.; Kinoshita, M.; Maruyama, S.-z.; Osaka, Y.-s.; Masuko, T.; Kakehi, K. Anal. Biochem. 2007, 363, 245-257. Yamada, K.; Hyodo, S.; Matsuno, Y. K.; Kinoshita, M.; Maruyama, S. Z.; Osaka, Y. S.; Casal, E.; Lee, Y. C.; Kakehi, K. Anal. Biochem. 2007, 371, 52-61). The method allows rapid release of O-glycans as the reducing form within 10 min. In the present study, we connected the device to a MALDI-TOF MS spotter and achieved routine analysis of O-glycans in biological samples for clinical use after in situ derivatization of the released O-glycans with phenylhydrazine. We applied the method to the analysis of O-glycans expressed on MKN45 cells derived from human stomach cancer cells and found that MKN45 cells expressed characteristic trisialo-polylactosamine-type glycans as reported previously (Yamada, K.; Kinoshita, M.; Hayakawa, T.; Nakaya, S.; Kakehi, K. J. Proteome Res. 2009, 8, 521-537). We also applied the method to the analysis of O-glycans in serum samples. The present technique is the first attempt to use MS measurement for routine clinical diagnostic works.

MeSH terms

  • Carbohydrate Sequence
  • Cell Line, Tumor
  • Chromatography, High Pressure Liquid
  • Glycoproteins / chemistry
  • Humans
  • Molecular Sequence Data
  • Mucins / chemistry
  • Phenylhydrazines / chemistry
  • Polysaccharides / blood*
  • Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / methods*

Substances

  • Glycoproteins
  • Mucins
  • Phenylhydrazines
  • Polysaccharides
  • phenylhydrazine