Figure 1: Alignments of peptides found on anti-FLAG® M1 (left) resp. anti-FLAG® M2 (right). The antibodies were raised against the DYKDDDDK-peptide. Anti-FLAG® M2 is considered to be more specific. Relative abundance of the amino acids surrounding the DYK motif explains the higher specificity of the anti-FLAG® M2 antibody. Data is based on 1,347 resp. 1,940 different DYK-sequences.

The examples on the left (Fig. 1) show fingerprints of the FLAG®-M1 and –M2 antibodies (Sigma) raised against the DYKDDDDK peptide. This analysis of all sequences containing DYK was carried out for the amino acids following this motif and results in the fingerprints of the observed amino acid distribution. The mostly preferred and more specific FLAG®-M2 antibody shows a significantly more specific fingerprint. (Data with permission from Fraunhofer IZI, Leipzig)

 

 

Figure 2: Fingerprint for CD227 mAb recognizing the Mucin-1 PDTRP motif. Variance of the central amino acid in all sequences containing PDxRP.

Figure 2 represents a fingerprint for the epitope PDTRP recognized by the Mucin-1 antibody. The statistical epitope fingerprinting data suggests that this antibody prefers other amino acids but Threonine in the central position. Other amino acids are enriched as well. So the recognized motif should be denominated as PD(T)RP instead. This would be in perfect agreement with the structural problem to recognize all five consecutive amino acids with the same precision. It is also indicating a potential contact in the antibody’s structure, which cannot be discovered with standard methods. A complete report for the Mucin-1 antibody is available here.