Km9002t

HEp-2 cells were infected with HSV-1 at an MOI of 0.01 and were incubated with or without R. tanguticum nanoparticles (350 µg/ml) at intervals of 6, 12, 18, and 24 h post-infection. Cells were harvested in RIPA Lysis Buffer (Biosharp, Hefei, China) and the soluble fraction was then clarified by centrifugation at 12,000 × g for 5 min at 4°C. Equal amounts of protein (40 µg/sample) were then isolated by 8% sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and transferred to a pre-equilibrated PVDF membrane (Thermo Scientific). Membranes were blocked with 5% BSA for 2 h, rinsed and followed by incubation with anti-ICP4 antibody (Abcam, ab6514, 1:1000), anti-ICP8 antibody (Abcam, ab20194, 1:500) and anti-GAPDH antibody (Tianjin Sungene Biotech KM9002T, Beijing, China, 1:5,000) in 5% BSA at 4°C overnight, respectively. The membranes were washed and incubated with horseradish peroxidase-conjugated secondary antibodies for 2 h at room temperature and visualized by using an ECL Western Blot Detection Kit (Millipore Corp., Bedford, MA).

Western blot analysis was performed according to previous description [27 (link)]. Total protein (30 μg protein per lane) from MSCs transfected with different miRNAs was subjected to SDS-PAGE and immunoblotting with the antibody against poly ADP-ribose polymerase (PARP) (46D11; CST), CD63 (ab108950; Abcam), Collagen I (14695; Proteintech), α-SMA (ab5694; Abcam), and GAPDH (KM9002T; Sungene Biotech).