Development of a lateral flow immuno-chromatic strip assay for the detection of cyprinid herpesvirus 3 (CyHV-3)

Cyprinid herpesvirus 3 (CyHV-3) is the main pathogen of koi herpesvirus disease (KHVD), which has caused serious damage to the ornamental and food-producing carp industry. Effective and rapid on-site detection methods are needed for early diagnosis of CyHV-3. A lateral flow immuno-chromatographic assay (LFIA) using two specific anti-CyHV-3 monoclonal antibodies has been developed and validated for on-site detection of CyHV-3. MAb 3C9 was used to bio-conjugate CyHV-3 antigen with colloidal gold, and MAb 2A8 was used to capture antigen bound colloidal gold on the test line. The control line was lined with goat anti-mouse IgG to capture unbound colloidal gold to validate performance. The test results can be viewed within 10 min after putting the strip into CyHV-3 virus infection fluid. The lowest limit of detection for the LFIA test was found to be 1.5 × 10⁴ copies/μL and it showed no cross-reactivity with other fish viral pathogens. The specificity of the strip was 100% when spleen and kidney tissues of CyHV-3-infected and healthy koi were validated at the field level. The LFIA strip will be an effective device for the early detection of CyHV-3 in the future.