Video showing the internalization of VLP-Wuhan:(E, S, N, M&M-mCherry&M-pHluorin) into VeroE6. This VLPs allow the simultaneous tracking of the pH of the VLP together with its membrane dynamics. pHluorin emits bright fluorescent light in pH≥8 but its intensity sharply decreases at pH<7.5, completely disappearing at pH<5.As the M protein C-terminal domain lies inside the VLPs, the fused pHluorin serves as a real-time indicator of the intra-VLP pH. When added to the medium (pH8), pHluorin-labelled VLPs emitted a bright green signal  and mCherry emits bright red signal which renders the visible signal as yellow. However, shortly after binding to the membrane of the VeroE6 cells, the intensity rapidly disappeared, due to drop into pH during internalization and only the mCherry (Red) one is still present.

VeroE6

VeroE6 cells were isolated from kidney epithelial cells extracted from an African green monkey (Chlorocebus sp.)

No cell labelling,

SARS-CoV-2 VLPs Wuhan

Wild Type SARS-CoV-2 virus-like particles containing all the structural proteins.

M-mCherry M-pHluorin, Part of the M protein of the particle is tagged with mCherry and part with M-pHluorin. This allows us to track simultaneously the membrane of the particle and the the change of the pH under the microscope.

No treatment