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Clinical and Diagnostic Laboratory Immunology, 11 1995, 652-656, Vol 2, No. 6
Copyright © 1995 by the American Society for Microbiology. All rights reserved.

Nucleocapsid protein N of Lelystad virus: expression by recombinant baculovirus, immunological properties, and suitability for detection of serum antibodies

JJ Meulenberg, RJ Bende, JM Pol, G Wensvoort and RJ Moormann
Department of Virology, Institute for Animal Science and Health (ID- DLO), Lelystad, The Netherlands. J.J.M.Meulenberg@id.agro.nl

The ORF7 gene, encoding the nucleocapsid protein N of Lelystad virus (LV), was inserted downstream of the P10 promoter into Autographa californica nuclear polyhedrosis virus (baculovirus). The resulting recombinant baculovirus, designated bac-ORF7, expressed a 15-kDa protein in insect cells. This protein was similar in size to the N protein expressed by LV in CL2621 cells when it was analyzed on sodium dodecyl sulfate-polyacrylamide gels. The N protein expressed by bac- ORF7 was immunoprecipitated with anti-ORF7 was immunoprecipitated with anti-ORF7 peptide serum, porcine convalescent-phase anti-LV serum, and N protein-specific monoclonal antibodies, indicating that this N protein had retained its native antigenic structure. The recombinant N protein was immunogenic in pigs, and the porcine antibodies raised against this protein recognized LV in an immunoperoxidase monolayer assay. However, pigs vaccinated twice with approximately 20 micrograms of N protein were not protected against a challenge with 10(5) 50% tissue culture infective doses of LV. Experimental and field sera directed against various European and North American isolates reacted with the N protein expressed by bac-ORF7 in a blocking enzyme-linked immunosorbent assay. Therefore, the recombinant N protein may be useful for developing diagnostic assays for the detection of serum antibodies directed against different isolates of LV.

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