The identification of b-lymphocyte epitopes of herpes simplex virus type 1 glycoprotein H.
thesisposted on 19.11.2015, 09:09 by James Steward. Chadwick
Herpes simplex virus type 1 glycoprotein H is a minor viral constituent which has been implicated in viral mechanisms of cell entry and exit. Study of this molecule has been hindered by a relative lack of specific immunological reagents. This study reports the generation and characterisation of specific polyclonal antibodies. The purification of gH and the generation of polyclonal serum is described. Antigenic determinants reactive with polyclonal antibodies were determined using synthetic peptides derived from the primary amino acid sequence of the molecule and a similar aproach was applied using available monoclonal antibodies. A series of 833 hexapeptides, with five residue overlap, were synthesised representing all potential continuous epitopes comprised of six residues or less. Peptides, displaying structural homology with epitopes of the native molecule were identified by screening for antibody binding. To further validate the identity of epitopes, corresponding homologous peptides were conjugated to carrier molecules and used to generate anti-peptide antibodies. The biological and immunological properties of the resulting anti-peptide antibodies were determined. Three antipeptide antibodies appeared reactive with the glycoprotein either following western transfer or in immunoprecipitation confirming the identity of gH-1 epitopes.