Next we sought to test the effect of a drug which is likely to influence the passage of vRNPs through the fusion pore. Nucleozin is a small molecule drug which is known to cause aggregation of vRNPs in cellulo or in vitro via bridging NPs together . Therefore, we hypothesized that influenza virus particles treated with nucleozin will contain aggregated vRNPs which will be less likely to pass through the fusion pores and therefore lead to higher dMG values. Influenza virus particles were incubated with nucleozin and then allowed to co-tether with vesicles to the SLB surface as described above. Interestingly, we measured a broader distribution of dMG values, and a higher mean value of 75.7±2.4 nm (Figure ) as compared to the case of fusion of an identical preparation of vesicles with an identical batch of untreated viruses (68.2±2.4 nm). The dMG's from viral fusion experiments along with those from dually labeled influenza virus (Figure ) were plotted into a cumulative distribution function (Figure ). Since 95% of the dMG's for the dually labeled influenza virus fall below 58 nm, we chose this number as a cut-off value for distinguishing between symmetric (<58 nm) and non-symmetric (>58 nm) modes of fusion. In the case of untreated sample, 39% of all dMG's fell below 58 nm while in case of nucleozin-treated sample, 30% of all dMG's fell below 58 nm (Figure ). The data supports our initial hypothesis that nucleozin causes aggregation of vRNPs and therefore transfers through the fusion pore less efficiently. Given that this analysis is subject to key assumptions such as choice of cut-off value for dMG, we suggest that the results should be interpreted in a semi-quantitative and relative sense rather than an absolute measure. Also, to derive meaningful conclusions, comparison between like systems should be made. Although nucleozin has been described to exert its anti-influenza action by causing nucleoprotein aggregation in the cytoplasm and thereby prevent its nuclear import , its effect on the endosomal escape has not been reported so far. Therefore, based on our prediction, it will be interesting to discover whether nucleozin and other nucleoproteinbinding drugs can be found to exert inhibitory effects on viral content release in cell-based studies as well.