Temperature-sensitive mutants of adenovirus type 5 have been isolated from virus stocks mutagenized by nitrous acid, hydroxylamine and 5-bromodeoxyuridine. The frequency of such mutants in the surviving fractions of nitrous acid and hydroxylamine-treated stocks was 8.4 and 9.6%, respectively, while it was around 0.55% in stocks of virus grown in 5-bromodeoxyuridine. Most of the mutants tested so far appear to be relatively stable and show little evidence of excessive leakiness or back mutation, and will be suitable for genetic analysis. Preliminary experiments show good complementation between four of these mutants, and they have been assigned to four complementation groups.