Adaptation de la méthode peptide-spot à la recherche de sites, de la protéine RNA2 du nodavirus, reconnus par différents sérums de loup (Dicentrarchus labrax).

Spot-Synthesis technique was applied to study protein frame or to develop antibodies with specific characteristics for use in diagnostic or research on human or mammals. In this work this technique was used to find sites on RNA2 protein of Nodavirus which are immunogenic for sea bass (Dicentrarchus labrax). It aimed to achieve synthetic peptide vaccine production against this virus. One hundred and nine overlapping peptides, of 15 amino acids frameshifted by three residues, representing the complete RNA2 protein sequence of Sb1 and Sb2 nodavirus were synthesized on a cellulose membrane by the Spot-Technique. The set of membrane bound peptides was probed by incubation with sea-bass or rabbit serum. Anti sea bass or anti rabbit alkaline phosphatase conjugated antibody revealed the binding. The membrane was further treated so as to remove precipitated dye and bound antibodies and reused when necessary.Serums of rabbit, sea bass bloodstocks and adult (100g) before an after injection with a live Nodavirus were tested. Serum from adult before and after injection of two peptides 15 amino acids including the N terminal for the former and the C terminal for the latter were also tested. In opposition with rabbit, the control fish serum bound aspecifically several peptide and strengthened the idea of low specificity of fish IgM.The first results permit to spot foreseeable difficulty resulting from adaptation of mammal’s technique to fish. Part of them should be resolved by technical adaptation as modification of peptide presentation or use of purified immunoglobulin rather whole serum but they comfirm that the immunogenicity of an epitope depend of the species.