We foresee that our method of probing viral content transfer to target vesicles using a fluorogenic dye will find many applications in virology and related disciplines. For example, viral infectivity is typically measured using cell culture-based methods which are typically expensive and time-consuming . Therefore alternate methods for quantification of viral infectivity which are rapid and do not require cell cultures are highly desirable. Cell-free measurement of viral infectivity may allow assessment of the effects of antiviral therapeutics like neutralizing antibodies or small molecules (48) which block cellular entry or inhibitors of fusion pore formation like IFITM3 in a straightforward manner. Cell-free methods will also be beneficial for measuring infectivities of BSL-3 agents like highly pathogenic avian influenza (HPAI) A strain H5N1 and SARS-CoV-2 rapidly while generating minimal biohazardous waste, which will facilitate pandemic preparedness research. In our method, detection of a content transfer event implies that a virus particle has functional spike proteins, contains nucleic acid, and forms a fusion pore, all of which are necessary conditions of infectivity. Due to the fluorogenic nature of the RNAbinding dye, detection of fusion events is essentially background-free, so counting of such events (i.e., new fluorescent spots) is straightforward and minimally prone to artifacts and spurious events. Also, our method is suitable for studying the fusion behavior of any enveloped virus given that the viral receptor-ligand pair is known. For instance, influenza viruses that bind to human receptors can be made to fuse with a target vesicle provided it is functionalized with α2,6 sialic acid containing glycolipids. Therefore, it will be interesting to investigate whether counting the number of full fusion events can be directly correlated to viral infectivity obtained from cell-based assays and such efforts are currently underway.