VSV virions, like those of many viruses, contain not only virus encoded-proteins, but also host proteins. Some of these proteins are enclosed within the virion structure, while others, in the case of enveloped viruses like VSV, are embedded in the host-derived membrane. While many of these incorporations could be “accidental”, others may reflect the mechanism of virus assembly, be necessary for viral function, impact host range and infection efficiency, or influence the outcome of subsequent infections. For example, a recent study showed that depletion of a number of host proteins normally incorporated into PF-4217903 herpes simplex virus type-1 virions not only reduced virion production in those cells, but that new infections with the resulting virions also yielded fewer progeny virions even in cells expressing normal levels of the host protein. The host-derived proteins of a virus may also affect the host immune response. This is an especially important consideration for viruses, including VSV, used in therapeutic applications where large numbers of virus particles are administered, as it may influence efficacy as well as the potential for adverse side effects. Incorporation of ICAM-I into the envelope of human immunodeficiency virus type-1 not only increased infection efficiency but also interfered with virus neutralization by host antibodies. In another example, the presence of host complement control proteins such as CD46, CD55 and CD59 in the viral envelope has been shown to protect against antibody dependent complement mediated virus lysis in several viruses including human T cell leukemia/ lymphoma virus type I, human cytomegalovirus, hepatitis C virus, HIV-1, extracellular enveloped vaccinia virus, simian virus 5 and mumps virus. To comprehensively look at host protein incorporation, a number of purified viruses have been analyzed by mass spectrometry including poxviruses, herpesviruses, orthomyxoviruses, coronaviruses, retroviruses, paramyxoviruses, baculoviruses, hytroviruses and arteriviruses. Our previous study examined VSV virions grown in three different cell lines originating from different species, and found a number of similarities and differences in the host protein content. Here we conduct an analysis not only of intact virions, but also virions treated with proteinase K to remove surface proteins to look at the localization of host protein incorporation into the virion. The most highly enriched clusters, when looking at all the proteins, were vesicles and vesicle mediated transport, protein localization and nucleotide binding.