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Veterinary research communications2004; 28(5); 437-446; doi: 10.1023/b:verc.0000034996.18533.90

Development of a neutralizing monoclonal antibody-based blocking ELISA for detection of equine herpesvirus 1 antibodies.

Abstract: A single-dilution, sensitive and specific monoclonal antibody-based blocking enzyme-linked immunosorbent assay (B-ELISA) was developed as an alternative to the cumbersome virus neutralization test (VNT) for detection of equine herpesvirus-1 (EHV-1) antibodies. Neutralizing monoclonal antibodies (1H6 and 9C6) raised against EHV-1 (Hisar-90-7 strain) and sera from 70 horses (30 known negative and 40 known positive for EHV-1 antibodies by VNT) were used for standardization of the B-ELISA. Using a single serum dilution of 1:250 in B-ELISA, 100% specificity was obtained with both monoclonal antibodies (Mabs) in comparison to VNT. Similarly, the sensitivity of the B-ELISA was 92.5% and 100% with 1H6 and 9C6 Mabs, respectively. A very high correlation coefficient (r = 0.85) was observed between B-ELISA and VNT that was significant at the p < 0.01 level. B-ELISA detected a more than 3-fold rise in antibody titres in paired serum samples collected from mares aborting owing to EHV-1 infection. Mab 9C6 was chosen for testing 231 field sera from apparently healthy vaccinated and non-vaccinated horses from organized breeding farms belonging to 11 Indian states, and from Bhutan, by B-ELISA and VNT. There was very good agreement between the results obtained by both VNT and B-ELISA (K = 0.9438). Of 231 field sera, 144 samples were negative for EHV- 1 antibodies by both VNT and B-ELISA and 81 were positive by both tests. Two samples negative by VNT were found positive in B-ELISA. On the other hand, four weakly positive samples in VNT (VN antibody titre 0.9 1.2 log10) were negative in B-ELISA. The Mab (9C6)-based B-ELISA was found to be a suitable alternative to VNT for screening large numbers of field sera and enabled confirmatory EHV-1 serodiagnosis.
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Publication Date: 2004-09-24 PubMed ID: 15379438DOI: 10.1023/b:verc.0000034996.18533.90Google Scholar: Lookup
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  • Journal Article
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  • Non-U.S. Gov't

Summary

This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.

Research on a novel diagnostic tool for equine herpesvirus-1 (EHV-1) has been conducted, resulting in a new enzyme-linked immunosorbent assay (B-ELISA). The assay is based on monoclonal antibody technology, offering a more streamlined and sensitive alternative to the standard virus neutralization test (VNT).

Method and Materials

  • Monoclonal antibodies (Mabs) were raised against EHV-1, after extracting the virus from horses with a known positive EHV-1 infection.
  • The serum from 70 different horses (30 known negative and 40 known positive) were used alongside the Mabs to calibrate the assay.

Results

  • The B-ELISA showed 100% specificity to EHV-1, meaning that it exclusively detected this virus, without being affected by the presence of other potential substances. This high specificity was achieved regardless of the Mab employed.
  • The sensitivity was 92.5% with one Mab, and 100% with the other, indicating the B-ELISA’s accurate detection capabilities.
  • The assay also enabled detection of increased antibody titres in serum samples connected to EHV-1-induced abortions in mares, providing valuable information about the infection timeline.

Comparison with Virus Neutralization Test (VNT)

  • A significant positive correlation (p < 0.01; r = 0.85) between the results from the B-ELISA and the VNT was found, implying that both tests are reliable and similarly effective in detecting EHV-1.
  • When testing field sera from healthy horses (vaccinated and non-vaccinated), the B-ELISA demonstrated a very good agreement with the VNT results, and even detected EHV-1 in two samples that tested negative via VNT.
  • However, four samples that tested weakly positive in VNT were negative in B-ELISA.

Conclusion

  • Given its accuracy, sensitivity, and simplicity, the monoclonal antibody based B-ELISA offers a viable alternative to the more labor-intensive VNT for detecting EHV-1.
  • The assay further allows for efficient screening of large serum samples, which can expedite serodiagnosis of EHV-1.

Cite This Article

APA
Singh BK, Ahuja S, Gulati BR. (2004). Development of a neutralizing monoclonal antibody-based blocking ELISA for detection of equine herpesvirus 1 antibodies. Vet Res Commun, 28(5), 437-446. https://doi.org/10.1023/b:verc.0000034996.18533.90

Publication

Veterinary research communications
ISSN: 0165-7380
NlmUniqueID: 8100520
Country: Switzerland
Language: English
Volume: 28
Issue: 5
Pages: 437-446

Researcher Affiliations

Singh, B K
  • National Research Centre on Equines, Sirsa Road, Hisar-125001 (Haryana), India. bksinghnrce@yahoo.com.in
Ahuja, S
    Gulati, B R

      MeSH Terms

      • Animals
      • Antibodies, Monoclonal
      • Antibodies, Viral / blood
      • Enzyme-Linked Immunosorbent Assay / methods
      • Enzyme-Linked Immunosorbent Assay / veterinary
      • Herpesvirus 1, Equid / immunology
      • Herpesvirus 1, Equid / isolation & purification
      • Horses
      • Neutralization Tests / veterinary
      • Skin / embryology

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      Citations

      This article has been cited 2 times.
      1. He F, Kiener TK, Lim XF, Tan Y, Raj KV, Tang M, Chow VT, Chen Q, Kwang J. Development of a sensitive and specific epitope-blocking ELISA for universal detection of antibodies to human enterovirus 71 strains. PLoS One 2013;8(1):e55517.
        doi: 10.1371/journal.pone.0055517pubmed: 23383215google scholar: lookup
      2. Prabakaran M, Ho HT, Prabhu N, Velumani S, Szyporta M, He F, Chan KP, Chen LM, Matsuoka Y, Donis RO, Kwang J. Development of epitope-blocking ELISA for universal detection of antibodies to human H5N1 influenza viruses. PLoS One 2009;4(2):e4566.
        doi: 10.1371/journal.pone.0004566pubmed: 19238211google scholar: lookup
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