Epitope mapping

Epitope mapping (English epitope mapping) describes the determination of epitopes on antigens. Among them are MHCI -bound epitopes, which are recognized by the T cell receptor on cytotoxic T cells, and MHC II -bound epitopes, which are recognized by CD4-positive T cells as well as soluble or membrane - bound antigens, epitopes which are recognized by antibodies from B cells. The epitope mapping is used to identify potential vaccines, and the development of therapeutic and diagnostic products.

Determination of discontinuous epitopes

Antibodies can recognize both continuous and discontinuous epitopes. The determination of both types of epitopes may be performed by X-ray crystal structure analysis or by nuclear magnetic resonance spectroscopy, more rarely by means of chemical cross-linking or an alanine scan.

Provision of continuous epitopes

Continuous epitopes of antibodies can be detected by ELISPOT or by synthetic peptides (eg PVDF or cellulose) previously immobilized spatially separated on a membrane. Are then detected by the addition of the antibody and some washing steps with a secondary antibody conjugate, antibody-binding epitopes analogous to the Western blot.

T cell epitopes are only continuous epitopes. During MHCI presented epitopes have a length of eight to eleven amino acids, MHCII - presented epitopes between thirteen and seventeen amino acids long. T cell epitopes of the cytotoxic and helper T - cells can be identified in flow cytometry by ELISPOT or by tetramer staining. In a protein to be characterized peptides are synthesized to the desired length, which cover the entire sequence of the protein. To epitopes not miss the edges of the peptides used usually is an overlap of the peptides by five amino acids ( at MHCI ) to eight (for MHCII ) to the adjacent amino acid sequence in the peptides.

In cytotoxic T cells stimulates the release of radioactive 53Chrom of chromium -containing and ( per assay ) were measured with an epitope - laden victim cells after the addition of cytotoxic T cells. Alternatively, the formation of perforin or granzyme B are measured.

Helper T cells secrete, depending on the type, for a contact with MHCII - presented antigens characteristic cytokines can be measured, instead of the chromium.

Forecast

A limited prediction of MHC binding peptides is offered among others by the program or by the SYFPEITHI IEDB. IEDB provides a current database described epitopes. These predictions apply only mostly, so an experimental verification of the epitopes must follow.