Nature Methods
- 4, 437 - 444 (2007)
Published online: 8 April 2007; | doi:10.1038/nmeth1035
Multiplexed analysis of glycan variation on native proteins captured by antibody microarraysSongming Chen1, Tom LaRoche1, 4, Darren Hamelinck1, 4, Derek Bergsma1, Dean Brenner2, Diane Simeone2, Randall E Brand3 & Brian B Haab11
Van Andel Research Institute, 333 Bostwick, Grand Rapids, Michigan 49503, USA. 2
University of Michigan Medical Center, 1500 E. Hospital Drive, Ann Arbor, Michigan 48109, USA. 3
Evanston Northwestern Healthcare, 2100 Pfingstein Rd, Glenview, Illinois 60025, USA. 4
Present addresses: Wayne State University Medical School, 540 E. Canfield, Detroit, MI 48201, USA (T.L.), McMaster University, 1280 Main Street West, Hamilton, Ontario, Canada L8S 4L8 (D.H.).
Correspondence should be addressed to Brian B Haab brian.haab@vai.org Carbohydrate post-translational modifications on proteins are important determinants of protein function in both normal and disease biology. We have developed a method to allow the efficient, multiplexed study of glycans on individual proteins from complex mixtures, using antibody microarray capture of multiple proteins followed by detection with lectins or glycan-binding antibodies. Chemical derivatization of the glycans on the spotted antibodies prevented lectin binding to those glycans. Multiple lectins could be used as detection probes, each targeting different glycan groups, to build up lectin binding profiles of captured proteins. By profiling both protein and glycan variation in multiple samples using parallel sandwich and glycan-detection assays, we found cancer-associated glycan alteration on the proteins MUC1 and CEA in the serum of pancreatic cancer patients. Antibody arrays for glycan detection are highly effective for profiling variation in specific glycans on multiple proteins and should be useful in diverse areas of glycobiology research.
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G1 reaction buffer (New England Biolabs)
