Development of an antigen-capture ELISA for the detection of equine influenza virus nucleoprotein.

This study details the development of an antigen-capture enzyme-linked immunosorbent assay (AC-ELISA) for detecting the equine influenza virus, verifying its efficacy and testing its specificity against other equine viruses.

Objective of the Study

• The primary aim of the research was to develop an effective AC-ELISA capable of identifying the presence of the equine influenza virus. This was executed using both monoclonal and polyclonal antibodies aimed against the A/equine/Xingjiang/2007 (H3N8) nucleoprotein (NP).

Research Methodology

• Immunoglobulin G antibodies were purified and utilized as capture or detector antibodies. These antibodies are integral to the antigen identification process, acting as the detecting agent against the presence of the virus.
• The specificity of the developed AC-ELISA was evaluated by testing it using a variety of equine viruses: EIV, equine herpesvirus 1 (EHV-1), equine herpesvirus 4 (EHV-4), equine arteritis virus (EAV), and Japanese encephalitis virus (JEV). This was conducted to assess whether the developed AC-ELISA displayed specificity in detecting only the equine influenza virus.

Research Findings

• The AC-ELISA proved to be effective, with only EIV specimens producing a strong signal. This indicates that the AC-ELISA successfully detected the EIV but not other similar viruses, demonstrating high specificity.
• The assay detected a minimal protein concentration of 50 ng/ml EIV in Nonidet P40-treated virus preparations, revealing its effectiveness.
• The AC-ELISA was capable of detecting the virus in nasal swabs of horses that had been experimentally infected between 3 to 7 days post-infection. This was confirmed using virus isolation and multi reverse transcription polymerase chain reaction, further validating the efficacy of the AC-ELISA.
• Both H3N8 and H7N7 EIV subtypes returned a positive result when using the AC-ELISA, suggesting that it is effective in detecting all subtypes of EIV.

Conclusion

• The development of the AC-ELISA for the equine influenza virus has proved successful, demonstrating high specificity for this virus and its subtypes. This research provides a positive contribution to virus detection methods, potentially aiding in faster response times to equine influenza infections.