To measure the viral content distribution within the fused volume as illustrated in Figure , ideally the viral fusion experiment needs to be carried out in a configuration where the virus and vesicle are placed side-by-side. Initially we attempted to obtain this information by carrying out fusion between virus particles with vesicles tethered to a PLL-PEG coated surface as illustrated in Figure . We obtain a distribution of dMG values ranging roughly between 0-150 nm with a mean of 59.9±2.2 nm (Figure )a value significantly higher than the corresponding dMG values (<30 nm) obtained for objects where two signals are perfectly overlapping as shown in Figure . The data clearly indicates that many fusion events were characterized by a nonsymmetric content distribution which is expected to yield larger dMG values as compared to symmetric content distribution. However, we must also consider that in this fusion configuration, from simple geometric arguments, there will be certain limiting conditions where a symmetric and non-symmetric fusion volume will be indistinguishable. For example, a virus bound near the polar regions of a vesicle (as schematically shown in Figure ) will lead to nearly identical dMG values for both symmetric and non-symmetric modes of fusions (Figure ). In such situations, the dMG values will be underestimated for non-symmetric fused volumes.