The development of a competitive PCR-ELISA for the detection of equine herpesvirus-1

Doyle, Sean and Daly, Paul (2003) The development of a competitive PCR-ELISA for the detection of equine herpesvirus-1. Journal of Virological Methods, 107 (2). pp. 237-244. ISSN 0166-0934

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Equine herpesvirus-1 (EHV-1) infection is of significant animal welfare and economic importance. Yet, no standardised molecular techniques are available for diagnosis or confirmation of viral infection. The purpose of this study was to develop a standardised and quantitative assay system for the reliable detection of EHV-1 infection which was capable of eliminating the likelihood of false negative results. A region within the EHV-1 glycoprotein B gene was amplified by polymerase chain reaction (PCR), cloned and subjected to site-directed mutagenesis to generate a control plasmid, amplifiable by identical primers to wild type EHV-1, yet capable of detection by an alternate dinitrophenylated oligonucleotide probe in a PCR/ELISA system. A competitive PCR/ELISA system which can control for the presence of PCR inhibitors and which is capable of detecting 63 genome equivalents of EHV-1 has been developed. EHV-1 presence in infected equine tissue and cell culture material was demonstrated using this system. The entire assay can be completed within one working day and facilitates multiple sample analysis. The availability of a robust, competitive PCR/ELISA system for the detection of EHV-1 will facilitate the rapid and sensitive detection of EHV-1 and offers the potential for eliminating the occurrence of abortion storms in stud farms.

Item Type: Article
Keywords: Fetal; Diagnosis; Oligonucleotides; Immunoassay; Molecular diagnostics;
Academic Unit: Faculty of Science and Engineering > Biology
Item ID: 165
Identification Number: 10.1016/S0166-0934(02)00252-5
Depositing User: Dr. Sean Doyle
Date Deposited: 22 Dec 2004
Journal or Publication Title: Journal of Virological Methods
Publisher: Elsevier Science B.V.
Refereed: Yes
Funders: Enterprise Ireland (EI)

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