Thirty-four (34) days after arrival at our facility, a recently imported rhesus macaque demonstrated a grade 4/5 reaction to intradermal testing with mammalian old tuberculin and a strong positive response in a serum interferon gamma (IFN-γ) stimulation assay (Primagam®). The affected animal was euthanized to prevent further exposure of the other rhesus in the quarantine room. Necropsy revealed enlarged, caseating mediastinal lymph nodes. Further analysis confirmed the presence of Mycobacterium tuberculosis complex organisms. Strict isolation measures were initiated and intensive testing of all animals in the affected room began immediately. After 13 weeks of additional testing, none of the animals in the room showed any positive response and all were released from quarantine. This case illustrates the importance of prolonged quarantine of non-human primates (NHP) and illustrates the usefulness of serology-based diagnostics as an adjunct to intradermal testing (molecular-based diagnostics typically refers to polymerase chain reaction, whereas this assay is really serology based, even though it is an in vitro IFN-γ stimulation assay). It also demonstrates that with proper isolation procedures, the spread of tuberculosis can be prevented in NHP facilities.