RESUMO
A repertoire of monoclonal antibodies was generated by immunization of mice with cancer-associated glycoprotein CA19.9, and two of them were selected as optimal capture and detecting counterparts for sandwich test system for detection of CA19.9. Fine epitope specificity of the antibodies was determined using printed glycan array, enzyme-linked immunosorbent assay, and inhibitory enzyme-linked immunosorbent assay. Unexpectedly, both immunoglobulins did not bind key epitope of CA19.9 glycoprotein, tetrasaccharide SiaLeA, as well as its defucosylated form sialyl LeC (known as CA-50 epitope). The antibodies were found to have different glycan-binding profiles; however, they recognized similar glycotopes with common motif Galß1-3GlcNAcß (LeC), thus resembling specificity of human natural cancer-associated anti-LeC antibodies. We propose that cancer-specific glycopeptide epitope includes Galß1-3GlcNAcß fragment of a glycoprotein O-chain in combination with proximal hydrophobic amino acid(s) of the polypeptide chain.
