Focused differential glycan analysis with the platform antibody-assisted lectin profiling for glycan-related biomarker verification

Mol Cell Proteomics. 2009 Jan;8(1):99-108. doi: 10.1074/mcp.M800308-MCP200. Epub 2008 Aug 11.


Protein glycosylation is a critical subject attracting increasing attention in the field of proteomics as it is expected to play a key role in the investigation of histological and diagnostic biomarkers. In this context, an enormous number of glycoproteins have now been nominated as disease-related biomarkers. However, there is no appropriate strategy in the current proteome platform to qualify such marker candidate molecules, which relates their specific expression to particular diseases. Here, we present a new practical system for focused differential glycan analysis in terms of antibody-assisted lectin profiling (ALP). In the developed procedure, (i) a target protein is enriched from clinic samples (e.g. tissue extracts, cell supernatants, or sera) by immunoprecipitation with a specific antibody recognizing a core protein moiety; (ii) the target glycoprotein is quantified by immunoblotting using the same antibody used in (i); and (iii) glycosylation difference is analyzed by means of antibody-overlay lectin microarray, an application technique of an emerging glycan profiling microarray. As model glycoproteins having either N-linked or O-linked glycans, prostate-specific antigen or podoplanin, respectively, were subjected to systematic ALP analysis. As a result, specific signals corresponding to the target glycoprotein glycans were obtained at a sub-picomole level with the aid of specific antibodies, whereby disease-specific or tissue-specific glycosylation changes could be observed in a rapid, reproducible, and high-throughput manner. Thus, the established system should provide a powerful pipeline in support of on-going efforts in glyco-biomarker discovery.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antibodies / immunology*
  • Biomarkers / analysis*
  • CHO Cells
  • Cell Line, Tumor
  • Cricetinae
  • Cricetulus
  • Glycosylation
  • Humans
  • Lectins / analysis*
  • Membrane Glycoproteins / metabolism
  • Mice
  • N-Acetylneuraminic Acid / metabolism
  • Platelet Aggregation
  • Polysaccharides / analysis*
  • Prostate-Specific Antigen / metabolism
  • Protein Array Analysis / methods*
  • Rats
  • Reproducibility of Results
  • Tissue Extracts


  • Antibodies
  • Biomarkers
  • Lectins
  • Membrane Glycoproteins
  • PDPN protein, human
  • Polysaccharides
  • Tissue Extracts
  • Prostate-Specific Antigen
  • N-Acetylneuraminic Acid