Clonality characterization of natural epitope-specific antibodies against the tumor-related antigen topoisomerase IIa by peptide chip and proteome analysis: a pilot study with colorectal carcinoma patient samples

Anal Bioanal Chem. 2012 Apr;403(1):227-38. doi: 10.1007/s00216-012-5781-5. Epub 2012 Feb 16.


Patient-specific sequential epitopes were identified by peptide chip analysis using 15mer peptides immobilized on glass slides that covered the topoisomerase IIa protein with a frameshift of five amino acids. Binding specificities of serum antibodies against sequential epitopes were confirmed as being mono-specific by peptide chip re-analysis of epitope-affinity-purified antibody pools. These results demonstrate that serum samples from colon carcinoma patients contain antibodies against sequential epitopes from the topoisomerase IIa antigen. Interactions of patients' antibodies with sequential epitopes displayed by peptides on glass surfaces may thus mirror disease-specific immune situations. Consequently, these data suggest epitope-antibody reactivities on peptide chips as potential diagnostic readouts of individual immune response characteristics, especially because monospecific antibodies can be interrogated. Subsequently, the clonality of the antibodies present in the mono-specific antibody pools was characterized by 2D gel electrophoresis. This analysis suggested that the affinity-purified antibodies were oligoclonal. Similarly to large-scale screening approaches for specific antigen-antibody interactions in order to improve disease diagnostic, we suggest that "protein-wide" screening for specific epitope-paratope interactions may help to develop novel assays for monitoring of personalized therapies, since individual properties of antigen-antibody interactions remain distinguishable.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Antibodies / chemistry
  • Antibodies / immunology*
  • Antigens, Neoplasm / immunology*
  • Blotting, Western
  • Cell Line, Tumor
  • Chromatography, Affinity
  • Colorectal Neoplasms / enzymology
  • Colorectal Neoplasms / immunology
  • Colorectal Neoplasms / metabolism*
  • DNA Topoisomerases, Type II / immunology*
  • DNA-Binding Proteins / immunology*
  • Electrophoresis, Polyacrylamide Gel
  • Enzyme-Linked Immunosorbent Assay
  • Epitopes / chemistry
  • Epitopes / immunology*
  • Humans
  • Molecular Sequence Data
  • Pilot Projects
  • Proteome*
  • Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization


  • Antibodies
  • Antigens, Neoplasm
  • DNA-Binding Proteins
  • Epitopes
  • Proteome
  • DNA Topoisomerases, Type II