Attachment: a recombinant VACV with Gaussia luciferase (LUC) fused to a core protein is used to measure the binding of virions at 4 °C by assaying cell-associated LUC activity. Hemifusion: R18-loaded virions are bound to target cells at 4 °C, shifted to 37 °C, and the dequenching of R18 dimers due to lipid mixing is measured by increased fluorescence. Core entry: a recombinant VACV that expresses firefly LUC under an early promoter, is used to infect cells and newly synthesized LUC is measured.
Modified from: Laliberte, J.P., Weisberg, A.S., Moss, B., 2011. PLoS Pathog. 7, e1002446.