|Topic:||Choosing the most suitable protein sequence for generating an anti-peptide serum .|
|Details:|| Choosing the most suitable protein sequence for generating an anti-peptide serum is the first and most crucial step in a successful immunization project.
The ideal peptide sequence should be unique to the target protein and located in an optimal position for epitope-antibody interaction. E.g. some anti-peptide antibodies might not recognize their epitope(s) when used for targeting the native protein in certain applications. In addition, technical requirements (e.g. suitability for synthesis, conjugation and predicted solubility for immunization) have to be considered.
Agrisera offers custom synthesized peptides produced by solid phase peptide synthesis (using Fmoc-Chemistry) including a terminal cysteine with the possibility for optional modifications (e.g. acetylation, biotinylation and fluorophore coupling). Peptides can be synthesized from immunograde purity (> 70 %, sufficient for generating or testing antibodies) to higher purity (+ 95 %, required in enzymological and biological activity studies and other applications). The purity is controlled by HPLC and mass spectrometric analysis (amino acid analysis and sequencing on request).
Our peptide services are available separately or in combination with our Antibody production services.
Standard design and synthesis service
Based on your submitted target protein sequence (provided as .txt-file or by accession number) we suggest a maximum number of 3 peptide sequences to consider as suitable for synthesis. The chosen sequence (up to 14 amino acids + terminal cysteine) is synthesized in a 15 mg immunograde purity scale without any further modifications. The delivery time is 4 weeks.
Important note: our standard service does not contain a detailed analysis of predicted cross-reactions to non-target proteins within the organism of interest. However, information provided on protein sequences (.txt-files or accession numbers) to which cross-reaction should be excluded will be considered. If possible, please specify additional information on regions that you would like to be targeted preferentially (or avoided), signal peptides, or any known target protein modifications (such as phosphorylation sites).
Coupling to carrier protein
Antigenicity of synthetic peptides is dramatically elevated by coupling to a carrier protein of higher mass as degradation by the host animal is diminished. In combination with standard peptide synthesis a fraction of the peptide (typically 2 mg) can be coupled to one of the following standard carrier proteins:
KLH - The keyhole limpet hemocyanin protein is a copper-containing protein found in arthropods and mollusca. Its molecular size (> 450 kDa) varies with its dissociation-states depending on the pH. Agrisera offers a polyclonal anti-KLH antibody.
BSA - The bovine serum albumin subunit of 67 kDa is a protein frequently used in immunoassays to block non-specific binding sites. It should not be used as carrier if future assays involve BSA (for instance as blocking agent). Agrisera offers a polyclonal anti-BSA antibody.
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