Hyphenated technique for releasing and MALDI MS analysis of O-glycans in mucin-type glycoprotein samples.
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.
Yamada
Keita
K
School of Pharmacy, Kinki University.
Hyodo
Satomi
S
Kinoshita
Mitsuhiro
M
Hayakawa
Takao
T
Kakehi
Kazuaki
K
D002240:Carbohydrate Sequence; D045744:Cell Line, Tumor; D002851:Chromatography, High Pressure Liquid; D006023:Glycoproteins / Q000737:chemistry; D006801:Humans; D008969:Molecular Sequence Data; D009077:Mucins / Q000737:chemistry; D010659:Phenylhydrazines / Q000737:chemistry; D011134:Polysaccharides / Q000097:blood*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*
D006023:Glycoproteins; D009077:Mucins; D010659:Phenylhydrazines; D011134:Polysaccharides; C030299:phenylhydrazine