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MHC Peptide Binding Assays

Each MHC allele has a distinct peptide binding motif which favours certain amino acids at particular points in a sequence, known as anchor residues.  Functional assays such as ELISpot or intracellular cytokine staining (ICS) can determine whether a cell responds to a particular peptide, but do not fully answer the question of which MHC allele is involved in a response.  Once potential epitopes have been identified by functional assays, it may be necessary to determine  to which allele a peptide is binding (MHC restriction).  This is especially relevant if these epitopes are to be used for immunotherapeutic purposes or as a biomarkers for immune monitoring.

If the peptide does not bind well, it is unlikely to be a strong epitope.  The ideal T cell epitope binds quickly to the MHC complex and remains stable in this complex for a long period of time.  Peptides with 'off-rates' of greater than 3 hours are more likely to be good T cell epitopes.

ProImmune REVEAL and ProVE® Rapid Epitope Discovery System

ProImmune REVEAL & ProVE® is a novel technology from ProImmune for analyzing proteins for potential T cell epitopes within a matter of weeks. The in vitro MHC peptide binding assay screens individual peptides, or peptide libraries, for their ability to stabilize the MHC complex for a wide range of both class I6 and class II7 alleles.  This unique approach will measure both the 'on-rate' and 'off-rate' of each peptide for multiple alleles providing a more in depth understanding of the binding kinetics.  In general, good T cell epitopes tend to have rapid on-rates and slow off-rates.  The MHC-peptide binding assay assigns each peptide a score relative to a known T cell epitope that has a borderline affinity to the allele of interest and in this way the most immunogenic peptides can be identified.

This strategy can narrow down potential candidate epitopes for further investigation and as the assays are cell free, they do not waste patient samples on testing peptides that are unlikely to be antigenic. The technology is widely applicable across many disease areas including all areas of cancer and infectious diseases. New CD4+ and CD8+ T cell epitopes identified with the help of this technology can be used as core building blocks for vaccine development or as targets for new immunotherapy.  Alternatively when applied to therapeutic proteins, the information from the report can be used in conjunction with cellular assays to measure the immunogenicity of proteins in order to avoid adverse reactions or neutralizing antibody titres.  The MHC-peptide binding assay can also identify peptides that bind a wide range of DR alleles, thus allowing a therapy to be targeted to particular ethnic populations expressing specific HLA molecules.

Peptides identified as positive in MHC-peptide binding assays, with good binding properties, can be further validated as T cell epitopes using ProImmune's Pro5® MHC Class I Pentamers or ProT2® MHC Class II tetramers to identify antigen specific CD8+ or CD4+ T cells respectively.



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