Fig. 4
Aβ1–42 induced EV-A71 aggregation and directly bound to VP1. a Purified EV-A71 virus was incubated with Aβ1–42 (20 µg/mL) at 4 °C for 60 min, and EV-A71 aggregation was analyzed by TEM. b SH-SY5Y or Vero cells were subjected to EV-A71 infection (MOI = 2.5) for 8 h, and the co-localization between Aβ1–42 (detected with a 488-conjugated antibody against Aβ1–42, green) and EV-A71 VP1 (Alexafluor594-conjugated antibody against VP1, red) was identified by confocal assay. Images were captured at 100 × magnification with a PE UltraVIEW VOX. c Aβ1–42 (20 µg/mL) was mixed with activated protein A/G magnetic beads overnight at 4 °C. Subsequently, the mixture was cultivated with purified EV-A71 virus (MOI = 1) at 4 °C for h. After cleaning the beads to remove nonbound viruses, the magnetic beads were subjected to precipitation with a magnet holder and assayed by Western blot with VP1 antibody. d Pre-virus attachment inhibition assay of Aβ1–42 showed comparable inhibition capacity against EV-A71 infections. Virus (MOI = 2.5) with or without Aβ1–42 medium incubated at 4 °C for 1 h, followed by infecting precooled cells at 4 °C for another 1 h. The quantity of cell-bound EV-A71 virus was identified by qRT-PCR. Experiments were independently repeated three times