Fig. 1.

Overview of the experimental workflow and identification of complete amino acid sequence of BaGs6. (A) Mouse ascites BaGs6 (IgM) antibody, reactive to Tn antigen and used in a number of publications as a cancer biomarker, was purified using affinity chromatography with immobilized Asialo-BSM which carries a high density of Tn antigen. From complete amino acid sequences, the recombinant Tn antigen-specific human IgG1, named Remab6, was purified and characterized. (B) Asialo-BSM beads were prepared by neuraminidase treatment of BSM beads. High-density Asialo-BSM beads were stained with ascites or anti-STn antibody confirmed by microscopy analysis. (C) BaGs6-containing mouse ascites were affinity-purified with Asialo-BSM beads. (D) Complete amino acid sequences were determined by proteomic de novo sequencing and CDRs of H chain (top) and L chain (bottom) were compared between three established anti-Tn antibodies (83D4, MLS128 and 5E5). * indicates identical residues in all four antibodies. Red boxes indicate different residues between BaGs6 and one or more of the antibodies.