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Recombinant nucleocapsid protein-based IgG enzyme-linked immunosorbent assay for the serological diagnosis of SARS.

ABSTRACT: The recombinant nucleocapsid protein (rNP) of severe acute respiratory syndrome (SARS) coronavirus (SARS-CoV) was expressed in a baculovirus system. The purified SARS-CoV rNP was used as an antigen for detection of SARS-CoV antibodies in IgG enzyme-linked immunosorbent assay (ELISA). The ELISA was evaluated in comparison with neutralizing antibody assay and the authentic SARS-CoV antigen-based IgG ELISA. Two-hundred and seventy-six serum samples were collected from health care workers in a hospital in which a nosocomial SARS outbreak took place and used for evaluation. The SARS-CoV rNP-based IgG ELISA has 92% of sensitivity and specificity compared with the neutralizing antibody assay and 94% sensitivity and specificity compared with the authentic SARS-CoV antigen-based IgG ELISA. The results suggest that the newly developed SARS-CoV rNP-based IgG ELISA is a valuable tool for the diagnosis and seroepidemiological study of SARS. The SARS-CoV rNP-based IgG ELISA has an advantage over the conventional IgG ELISA in that the antigen can be prepared by laboratory workers without the risk of infection.

SUBMITTER: Saijo M

PROVIDER: S-EPMC7112814 | biostudies-literature | 2005 May

REPOSITORIES: biostudies-literature

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Recombinant nucleocapsid protein-based IgG enzyme-linked immunosorbent assay for the serological diagnosis of SARS.

Saijo Masayuki M Ogino Toshio T Taguchi Fumihiro F Fukushi Shuetsu S Mizutani Tetsuya T Notomi Tsugunori T Kanda Hidetoshi H Minekawa Harumi H Matsuyama Shutoku S Long Hoang Thuy HT Hanh Nguyen Thi Hong NT Kurane Ichiro I Tashiro Masato M Morikawa Shigeru S

Journal of virological methods 20050501 2

The recombinant nucleocapsid protein (rNP) of severe acute respiratory syndrome (SARS) coronavirus (SARS-CoV) was expressed in a baculovirus system. The purified SARS-CoV rNP was used as an antigen for detection of SARS-CoV antibodies in IgG enzyme-linked immunosorbent assay (ELISA). The ELISA was evaluated in comparison with neutralizing antibody assay and the authentic SARS-CoV antigen-based IgG ELISA. Two-hundred and seventy-six serum samples were collected from health care workers in a hospi ...[more]

PMID: 15794988

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Project description:BackgroundSchistosomiasis is a huge threat to human and animal health. Apart from bovines, goats play an important role in the transmission of schistosomiasis in some endemic areas of China. An accessible, quality-assured goat schistosomiasis diagnostic technique is needed. Recently, our laboratory identified two recombinant diagnostic antigens, SjPGM and SjRAD23 via an immuno-proteomic method. The application of these two recombinant antigens to develop a higher sensitivity and specificity technique for the sheep schistosomiasis diagnosis is urgently needed.MethodsEpitopes of SjPGM and SjRAD23 were predicted and three polypeptides, two from SjRAD23 and one from SjPGM, were selected. Recombinant plasmids containing two to three DNA sequences encoding predicted polypeptides or large hydrophilic region of Sj23 (LHD-Sj23) were constructed and expressed. Eight recombinant schistosome antigens including four multi-epitope proteins and four recombinant single-molecule antigens as well as SEA, were assessed by ELISA in 91 sera from schistosome-infected goats, 44 sera from non-infected goats, 37 sera from Orientobilharzia-infected goats, and 12 from Haemonchus contortus-infected goats.ResultsELISA tests showed that three multi-epitope proteins had higher sensitivity than the four single-molecule antigens (rSjRAD23, rSjPGM, rBSjRAD23-1, rBSj23) and the multi-epitope protein rBSjPGM-BSjRAD23-1-BSj23 had the highest sensitivity (97.8 %, 89/91) and maintained good specificity (100 %, 44/44) as well as low cross-reactivity with haemonchosis (8.33 %, 3/12) and orientobilharziasis (13.51 %, 5/37) in the diagnosis of goat schistosomiasis. In contrast, when SEA was applied as a diagnosis antigen, it had 100 % (91/91) sensitivity, 75 % (33/44) specificity, 25 and 83.78 % cross-reactivity with haemonchosis (3/12) and orientobilharziasis (31/37), respectively.ConclusionsThe application of recombinant multi-epitope proteins may increase the sensitivity of diagnosis technique and retain high specificity of single-molecule antigens for schistosomiasis, and the recombinant antigen rBSjPGM-BSjRAD23-1-BSj23 has the potential to be used as a diagnosis antigen for goat schistosomiasis.

Project description:Ovine enzootic abortion (OEA) resulting from infection of sheep and goats with Chlamydophila abortus is of major economic importance worldwide. Over the last 50 years the serological diagnosis of infection has been based mainly on the complement fixation test (CFT), which lacks both sensitivity and specificity because of cross-reactive antibodies to other gram-negative bacteria, including another common chlamydial pathogen of sheep, Chlamydophila pecorum. In the present study, a series of overlapping recombinant antigens representing the polymorphic outer membrane protein POMP90 of C. abortus was assessed by enzyme-linked immunosorbent assay (ELISA) with a panel of 143 serum samples from sheep experimentally infected with C. abortus, from sheep clinically free of OEA, and from specific-pathogen-free lambs experimentally infected with different subtypes of C. pecorum. The results were compared to those obtained by CFT and another recently described test, an indirect ELISA (iELISA) with the recombinant OMP91B (rOMP91B) fragment (rOMP91B iELISA) (D. Longbottom, E. Psarrou, M. Livingstone, and E. Vretou, FEMS Microbiol. Lett. 195:157-161, 2001). The rOMP90-3 and rOMP90-4 ELISAs were identified as being more sensitive and specific than CFT. Assays with both fragments were evaluated further with a panel of 294 field serum samples from flocks with documented histories of abortion, from flocks with no clinical histories of abortion but which had a high proportion of samples seropositive by CFT, and from animals with no histories of abortion but from which various C. pecorum subtypes had been isolated. ELISAs with both POMP90 fragments outperformed CFT with serum samples from C. pecorum-infected animals, producing no false-positive results. However, the ELISA with the rOMP90-4 fragment appeared to be more sensitive than the one with rOMP90-3, as it identified more of the OEA-positive samples. The ELISA with the rOMP90-4 fragment was also able to identify apparently healthy animals that were infected with an enteric strain of C. abortus in flocks that were probably infected with both enteric C. abortus and C. pecorum strains. The identification of animals infected with enteric C. abortus is extremely important in controlling the spread of OEA. Overall, the new rOMP90-4 ELISA was found to be a more sensitive and specific test than CFT for differentiating animals infected with C. abortus from those infected with C. pecorum.

Dataset Information

Recombinant nucleocapsid protein-based IgG enzyme-linked immunosorbent assay for the serological diagnosis of SARS.

Publications

Recombinant nucleocapsid protein-based IgG enzyme-linked immunosorbent assay for the serological diagnosis of SARS.

Similar Datasets

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