Characterization of Neospora caninum surface protein NcSRS2 based on baculovirus expression system and its application for serodiagnosis of Neospora infection

Y. Nishikawa, Y. Kousaka, K. Tragoolpua, X. Xuan, L. Makala, K. Fujisaki, T. Mikami, H. Nagasawa

Research output: Contribution to journalArticle

32 Citations (Scopus)

Abstract

The baculovirus expression system has proved to be a useful tool for the production of recombinant proteins. Here we have characterized the Neospora caninum surface protein NcSRS2 produced by two types of the recombinant virus and also have developed an enzyme-linked immunosorbent assay (ELISA) using recombinant NcSRS2 for the serologic diagnosis of Neospora infection. Western blot analysis showed two major protein bands that were detectable in insect cells infected with each recombinant baculovirus, and a lower-molecular-weight protein was detected in culture supernatants from a cell infected with the recombinant virus lacking the hydrophobic C-terminal tail. Analysis of the N-terminal amino acids showed that the secreted NcSRS2 lacked 6 kDa of the N-terminal signal peptide. Moreover, the detergent-soluble protein of insect cells infected with the recombinant baculovirus expressing the full-length NcSRS2 gene was used to develop an ELISA system based on specificity and reactivity to antisera against Toxoplasma gondii, Hammondia heydorni, or N. caninum. Anti-N. caninum mouse, dog, and bovine sera recognized the recombinant NcSRS2 on Western blots. Furthermore, we have shown that the developed ELISA system consistently discriminates indirect fluorescent-antibody test (IFAT)-positive bovine sera against N. caninum from IFAT-negative sera. These results indicate that the ELISA using baculovirus-expressed NcSRS2 can be useful for effective and reliable serodiagnosis of N. caninum infection.

Original languageEnglish (US)
Pages (from-to)3987-3991
Number of pages5
JournalJournal of clinical microbiology
Volume39
Issue number11
DOIs
StatePublished - Nov 14 2001

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Neospora
Baculoviridae
Serologic Tests
Membrane Proteins
Infection
Enzyme-Linked Immunosorbent Assay
Western Blotting
Serum
Insect Proteins
Viruses
Antibodies
Toxoplasma
Protein Sorting Signals
Recombinant Proteins
Detergents
Insects
Tail
Immune Sera
Proteins
Molecular Weight

ASJC Scopus subject areas

  • Microbiology (medical)

Cite this

Characterization of Neospora caninum surface protein NcSRS2 based on baculovirus expression system and its application for serodiagnosis of Neospora infection. / Nishikawa, Y.; Kousaka, Y.; Tragoolpua, K.; Xuan, X.; Makala, L.; Fujisaki, K.; Mikami, T.; Nagasawa, H.

In: Journal of clinical microbiology, Vol. 39, No. 11, 14.11.2001, p. 3987-3991.

Research output: Contribution to journalArticle

Nishikawa, Y. ; Kousaka, Y. ; Tragoolpua, K. ; Xuan, X. ; Makala, L. ; Fujisaki, K. ; Mikami, T. ; Nagasawa, H. / Characterization of Neospora caninum surface protein NcSRS2 based on baculovirus expression system and its application for serodiagnosis of Neospora infection. In: Journal of clinical microbiology. 2001 ; Vol. 39, No. 11. pp. 3987-3991.
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