In general, a good peptide antigen has the following properties: protein surface location, flexible (usually loop) rather than helical structure, complicate and unique sequence, not a post-translational modification site or functional site (unless the antibody is intended to recognize such a site) and easy for synthesis. But the quantitative relationship between these properties and antigenic strength is not clear
Peptide antigen design is usually done by two different approaches: predicting peptide’s physical-chemistry properties or making prediction based on statistic results. Both approaches have their limitations. Physical chemistry properties such as peptide location in a protein or its secondary structure, particular turn, are difficult to be predicted with high degree of accuracy. This is because the problem themselves are parts of one of the most challenge areas of modern science —- protein folding. Another problem is that in most cases, an isolated peptide in solution can not maintain its native conformation found in the protein. This is the major reason that antigens designed based on known 3D structures are also often failed.
Although Cys residues are often added to peptides to enable crosslinking to a carrier protein, a peptide synthesized with many Cys residues present can be difficult to handle and may not lead to a useful antibody. Multiple Cys residues may lead to the formation of covalently linked aggregates. The Cys-rich regions of proteins may have some disulfide bonds. The linear peptide with reduced Cys would therefore not represent the protein itself, which would be more structurally constrained.
Most peptide antigen requested range in length from 12 to 20 residues and are relatively easy to synthesize. No Cys residues should be internal to the peptide sequence.