Adenovirus 8 (Ad 8, strain 1127) has the general structural features of an adenovirus; it produces the genus-specific hexon antigen. The fiber length of 12 nm corresponds to that of other subgenus D adenoviruses. In hemagglutination-inhibition, Ad 8 is related to Ad9 and Ad 15/H9. The slow multiplication, compared to most other subgenus D viruses, was shown not to be caused by an extended growth cycle, but appears to be due to an inefficient virus release into the medium. After restriction analysis with the enzymes Sma I and Bgl II, the typical fragment patterns of species from subgenus D were obtained, while very different patterns emerged from the analysis with Hind III, Bam HI, and Bst E II endonucleases. Several strains of Ad 8 produced more penton-associated toxin than Ad 15/H9 used as reference virus from subgenus D, which could explain the "clumping CPE" caused by Ad 8. Ad 8 toxin was neutralized by antisera from many adenovirus species. The poor virus yield of Ad 8 was not improved by removing soluble viral or cellular substances; all other attempts to improve virus yield by varying the conditions of cell culture also failed.