Development of a loop-mediated isothermal amplification for visual detection of the HCLV vaccine against classical swine fever in China.

A loop-mediated isothermal amplification (LAMP) assay was developed and evaluated for the rapid and specific detection of HCLV vaccine strain against classical swine fever. Four primers were designed for amplification of NS5B gene region with Bst DNA polymerase at a constant temperature of 65°C. The products showed ladder-like pattern on 2% agarose gel, and can be visualised after addition of SYBR Green I dye. The detection limit of the assay was 5 copies of the HCLV genome per reaction. No cross-reaction with other porcine viruses including different wild-type CSFV strains and the bovine viral diarrhoea virus was observed. The agreement between the LAMP and TaqMan real-time RT-PCR assays was 94.4% for the detection of 72 batches of HCLV vaccine. The assay provides a rapid tool for the control of vaccine quality and can be an accompanying assay of the LAMP for wild-type CSFV described previously for differential diagnosis.