Immunogen design

We can provide assistance with the design of appropriate peptide immunogens taking into account protein homology, antigenicity, hydrophilicity, accessibility and synthesis suitability by analysing complete or partial protein and/or gene sequences.

We will outline peptide regions of approximately 10-20 amino acids that are antigenic, hydrophilic, flexible and accessible on the protein surface. This will increase the likelihood of generating antibody capable of recognising the native protein. Although shorter sequences may be more specific, they will more likely be not as accessible on the protein surface. Longer sequences, on the other hand maybe less specific but more likely to produce antibodies recognising the native protein. Peptide between 10 and 20 amino acid long will have some secondary structure and will be more soluble.

It is essential that, when choosing a peptide, solubility, flexibility and surface exposure on the protein are taken into account. Hydrophilic sequences are normally exposed on the surface of the protein in contact with the water environment while hydrophobic residues tend to be buried in the middle of the structure away from water. Therefore, antibodies will be raised against hydrophilic epitopes that are easily accessible on the surface of the protein. It has also been shown that most antigenic sequences have a certain degree of flexibility within the overall structure of the protein.

Another pointer in choosing a good candidate peptide sequence is to look at the C- and N-terminus sequences. These are normally on the surface of the protein and flexible, making them ideal candidates. However, if they contain hydrophobic residues, then it is probable that they are not exposed at all. Also, make sure to avoid any region that might be included in the transmembrane sequences of surface receptors since, obviously, these regions will not be accessible to antibodies.

All peptides will undergo a BLAST search to confirm specificity and rule out unwanted antibody crossreactivity with related or unrelated proteins. This is extremely important since short peptides sequences may show high homology to sequences in other proteins. We will pass the information to the researcher who will decide whether potential crossreactions might be problematic or not.

If there are 2 or more related proteins, a sequence alignment analysis will also be performed. This is essential to avoid cross-reaction between different protein by choosing unique sequences. This will also identify amino acid homology and show the position of conserved regions.

If you wish us to analyse your protein sequence and suggest good potential peptide, send us the DNA or the protein sequence and provide the name of the species involved and the name and accession number of the protein(s) in question. Please provide the amino acid sequence using the single letter code. If the protein sequence is derived from a cDNA sequence, it is essential not to use the leader sequence in the analysis to generate an antibody.

If you already have it, send us the sequence alignment of all related proteins to help us avoiding regions of conserved amino acids. Also, let us know if you have any preference for any region (amino, carboxyl end, etc.).