Identification of canine helper T-cell epitopes from the fusion protein ofcanine distemper virus

The fusion protein of canine distemper virus (CDV-F), a 662 amino-acid enve lope protein, was used as the target molecule for identification of canine T helper (Th) epitopes. A library of 94 peptides, each 17 residues in lengt h overlapping by 10 residues and covering the entire sequence of CDV-F, was screened using a lymphocyte proliferation assay with peripheral blood mono nuclear cells (PBMC) obtained from dogs inoculated with canine distemper vi rus (CDV) vaccine. Initially we observed low and inconsistent proliferation of PBMC in response to these peptides, even when using cells obtained from dogs that had received multiple doses of CDV. Subsequently, the use of exp anded cell populations derived by in vitro stimulation of canine PBMC with pools of peptides allowed the identification of a number of putative canine Th-epitopes within the protein sequence of CDV-F. There were two major clu sters of Th-epitopes identified close to the cleavage site of the F0 fusion protein, while some others were scattered in both the F1 and F2 fragments of the protein. Some of these peptides, in particular peptide 35 (p35), wer e stimulatory in dogs of different breeds and ages. The identification of s uch promiscuous canine Th-epitopes encouraged us to assemble p35 in tandem with luteinising hormone releasing hormone (LHRH) a 10 amino-acid residue s ynthetic peptide representing a B-cell epitope which alone induces no antib ody in dogs, The totally synthetic immunogen was able to induce the product ion of very high titres of antibodies against LHRH in all dogs tested. Thes e results indicate that p35 could be an ideal candidate for use as a Th-epi tope for use in outbred dogs.