Viruses package their genome in a robust protein coat to protect it during transmission between cells and organisms. In a reaction termed uncoating, the virus is progressively weakened during entry into cells. At the end of the uncoating process the genome separates, becomes transcriptionally active, and initiates the production of progeny. Here, we triggered the disruption of single human adenovirus capsids with atomic force microscopy and followed genome exposure by single-molecule fluorescence microscopy. This method allowed the comparison of immature (noninfectious) and mature (infectious) adenovirus particles. We observed two condensation states of the fluorescently labeled genome, a feature of the virus that may be related to infectivity. Beyond tracking the unpacking of virus genomes, this approach may find application in testing the cargo release of bioinspired delivery vehicles.