Table 2.1.

Comparison of single virion and ensemble methods for studying particular viral entry steps, including key features of each method

Virus entry step	Single virion tracking		Ensemble methods
	Live cell	Biomimetic
Extracellular transport	• Direct cell-cell spread • Extracellular diffusion	• Movement through mucosa
Cell-surface trafficking	• Cytoskeletal interaction • Movement toward entry site	• Diffusion, rolling, and rocking along surface • Bilayer composition • Bilayer fluidity
Binding	• Colocalization with receptor	• Attachment/detachment rates • Bilayer composition • Bilayer fluidity • Receptor mobility • Adhesion-strengthening	QCMD Coflotation ELISA SPR TEM
Internalization	• Clathrin dependence/independence • Internalization timescale • Cytoskeletal interaction	n/a	IFA TEM
Fusion	• Differentiate plasma membrane fusion from endosomal fusion • Escape from early vs. late endosomes	• Bilayer composition • Bilayer fluidity • Viral fusion environment • Timing/sequence of fusion triggers • Hemifusion and pore formation rate constants • Number of rate-limiting fusion steps • Acid stability	TEM Syncytia formation Bulk solution fluorescence Infectivity BlaM release
Intracellular trafficking	• Cytoskeletal interaction • Extra- and intra-nuclear movement	n/a	IFA

Acronyms: QCMD Quartz crystal microbalance with dissipation, ELISA Enzyme-linked immunosorbent assay, SPR Surface plasmon resonance, TEM Transmission electron microscopy, IFA immunofluoresence assay, BlaM Beta lactamase