Porcine IL-12 plasmid as an adjuvant improves the cellular and humoral immune responses of DNA vaccine targeting transmissible gastroenteritis virus spike gene in a mouse model

LI, Xunliang; LI, Pengchong; CAO, Liyan; BAI, Yunyun; CHEN, Huijie; LIU, He; REN, Xiaofeng; LI, Guangxing

A lactate dehydrogenase (LDH) test kit was used to analyze cytotoxicity. The test was conducted according to the manufacturer's instructions (Jiancheng, Nanjing, Jiangsu, China). The effector cells were obtained from the blood and spleen lymphocytes of 42 dpi mice [4, 8] , and the target ST cells were infected with TGEV (70 PFU) for 36 hr (37°C, 5% CO 2 ). The effector cells and target cells were mixed at a proportion of 25:1 and incubated in wells of 96-well plates for 6 hr (37°C, 5% CO 2 ). The supernatants (100 µl) were mixed with LDH assay reagent (100 µl/well) for 15 min (37°C) in 96-well plates and the results were analyzed at 490 nm wavelength. The maximum release of LDH was that the total of target cells were dissolved in Triton X-100 (1% v/v), and the spontaneous LDH release was that the target cells exist single. Cytotoxic T lymphocytes (CTLs) were calculated as ([experimental release−spontaneous release]/[maximum release−spontaneous release]) ×100% [3] . All detections were performed in triplicates.