Polymerase chain reaction combined with fluorescent lateral flow immunoassay based on magnetic purification for rapid detection of canine parvovirus 2

Zhuang, Linlin; Ji, Yongxin; Tian, Peilong; Wang, Kaixuan; Kou, Chengkun; Gu, Ning; Zhang, Yu

To determine the sensitivity of CPV-2 detection by PCR-GE and PCR-LFIA, the standard DNA stock (3 × 10 8 copies/μL) was serially diluted tenfold. Each viral DNA (1 μL) was used as a template for PCR where the respective dilutions were subjected to thermal cycling using the primer pair of biotin-labeled CPV-2-F and Digoxigenin-labeled CPV-2-R, which amplified a 253-bp fragment from the VP2 gene of CPV-2. In PCR-LFIA, standard DNA dilutions gave a fluorescence signal values ranging from 131 to 23,251, and a linear relationship (y = 1577.8Ln(x)-7869.4, R 2 = 0.9675) was observed with CPV-2 titers decreasing from 3 × 10 8 to 3 × 10 0 copies/μL. The results showed that the sensitivity of PCR-LFIA for the detection of CPV-2 was 3 × 10 1 copies/ μL, which was 100 times more sensitive than PCR-GE assay ( Fig. 5 ; Table 3 ).