Background: Tigecycline is a minocycline derivative, belonging to a chemical class prone to oxidation and to medium-related effects on susceptibility. We therefore studied the effect of medium type and age on the tigecycline MICs found in broth and on agar by the Clinical Laboratory Standards Institute and British Society for Applied Chemotherapy methods. Methods: MICs of tigecycline, minocycline and tetracycline were determined for 96 non-fastidious bacteria and 20 streptococci using Mueller-Hinton broth (MHB), Mueller-Hinton agar (MHA), Iso-Sensitest broth (ISB) and Iso-Sensitest agar (ISA) as: (i) freshly-prepared media; (ii) fresh media but with the antibiotic added 1 day before inoculation and (iii) media stored for 7 days before antibiotic addition and use. Results: MICs of tigecycline in fresh MHB were up to two doubling dilutions higher than on or in MHA, ISA or ISB. Media with tigecycline or classical tetracyclines added a day before use gave raised MICs, though rarely by more than one dilution. MICs of tigecycline (less so classical tetracyclines) were increased in 7-day-old MHB or ISB, even though the antibiotic was freshly added. This latter effect was greatest for the most susceptible strains and was absent or much reduced for organisms with MICs ≥8 mg/L; it did not occur in the corresponding agar dilution tests. Addition of blood to MHB largely abrogated the effect, as did steaming the broth before adding the antibiotic. Conclusions: The raised MICs of tigecycline in aged broth probably reflect inactivation by dissolved oxygen. This accords with the lack of any MIC increase in newly-steamed (i.e. degassed) MHB or on aged agar (which is melted at 100°C before use). Blood, which also abrogated the effect, may increase reducing capacity, protecting the tigecycline. At a practical level, broth MIC determinations for tigecycline should always employ fresh media.