In our recombinant baculovirus system, VP1 protein of merkel cell polyomavirus (MCPyV), which is implicated as a causative agent in Merkel cell carcinoma, was self-assembled into MCPyV-like particles (MCPyV-LP) with two different sizes in insect cells, followed by being released into the culture medium. DNA molecules of 1.5- to 5-kb, which were derived from host insect cells, were packaged in large, ~50-nm spherical particles but not in small, ~25-nm particles. Structure reconstruction using cryo-electron microscopy showed that large MCPyV-LPs are composed of 72 pentameric capsomeres arranged in a T = 7 icosahedral surface lattice and are 48 nm in diameter. The MCPyV-LPs did not share antigenic determinants with BK- and JC viruses (BKPyV and JCPyV). The VLP-based enzyme immunoassay was applied to investigate age-specific prevalence of MCPyV infection in the general Japanese population aged 1-70 years. While seroprevalence of MCPyV increased with age in children and young individuals, its seropositivity in each age group was lower compared with BKPyV and JCPyV.