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How to select Antigenic Peptides from whole Protein Sequence and confirm their specificity using BLAST homology searches
Send the complete protein sequences or Gene Accession # to GSI and tell us where you would like to target the antibodies, if any preference, based upon conservation of sequence among various species or isoforms. We will find antigenic peptide based upon requirement.
GSI Inc. has been providing FREE consultation service (if you order services from GSI based) to Scientists around the globe to find specific antigenic peptides from long protein sequences. We realize that many researchers either do not have necessary tools, computers programs, and expertise or simply do not wish to invest time in analyzing protein sequences. We provide the following analyses.
1. Identification of specific antigenic peptides. Analyses of complete or partial protein sequences to find antigenic peptides. Our recommendations are based upon the analyses of antigenicity, hydrophilicity, and accessibility parameters. We try to find peptide regions (10-25 aa) that are antigenic, hydrophilic, and accessible. This information is sent in an easy to understand graphical format by email or fax. This increases the likelihood of generating an antibody that recognizes the full length protein.
2. BLAST searches. All recommended antigenic peptides are then subjected to BLAST to confirm specificity of peptides. This is extremely important since short peptides may show higher degree of amino acid homology with either related members or unrelated proteins creating unwanted antibody crossreactivity. It is very important to carefully review this information to avoid antibody reactivity to unwanted proteins fro the same or different species.
3. GSI may also analyze the protein secondary structure (see section 4). Based upon the available information, we typically suggest 1-4 peptides (10-25 AA) that are suitable for antibody production.
4. Proteins Secondary Structure Analyses. A given protein may contains one or more of the following.
- Signal peptide (GvH: von Heijne’s method for signal seq. Recognition)
- Transmembrane domains (ALOM: Klein et al’s method for TM region allocation)
- Mitochondrial targeting seq. (MITDISC)
- Nuclear localization signals (NUDISC)
- ER retention motif (KDEL)
- Peroxisomal targeting signal in the C-terminus (SKL/SKL2)
- Vacuolar targeting motif (VAC)
- Actinin-type actin-binding motif
- N-myristoylation pattern (NMYR)
- Prenylation motif
- Transport motif from cell surface to Golgi (memYORL)
- Tyrosines in the tail and Dileucine motif
- PROSITE DNA binding motifs Leucine zipper pattern
- PROSITE ribosomal protein motifs
- Reinhardt’s method for Cytplasmic/Nuclear discrimination
- Lupas’s algorithm to detect coiled-coil region
This information is very important in identifying the antigenic peptide and targeting antibodies to a desired domain. This info also helps in identifying the possible functions of proteins.
5. Multiple Sequence alignment using ClustalW . It is sometime necessary to produce multiple sequence alignments for 2 or more related proteins. It helps in identifying specific antigenic peptides and avoiding potential antibody crossreactivity.
How to submit protein sequence to us for analyses
Please send your name, complete address, phone, fax, email addresses preferably by email or fax. We will send our analyses by email or fax within 1-3 days. All information provided to us remains confidential. If you wish to submit a sequence for the identification of antigenic peptides for making antibodies, please provide the following information by email. Please do not forget to include your phone and fax #.
1. Name(s) of the protein, gene accession # (if available), and amino acid sequence (single letter code) by email.
2. Sequence alignment of all related proteins, if available, by fax. This will help us in avoiding regions that are conserved.
3. Any preference for targeting peptides (amino terminal, c-terminal). You can also specify regions to avoid.
4. You can mention any peptide(s) that you have already selected.
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