Animal olfactory detection of human diseases has attracted an increasing amount of interest from researchers in recent years. Because of the inconsistent findings reported in this body of research and the complexity of scent-detection research, it is difficult to ascertain the potential value of animal detectors in operational diagnostic algorithms. We have outlined key factors associated with successful training and evaluation of animals for operational disease detection and, using these key factors as points for comparison, conducted a systematic review of the research in this area. Studies that were published in peer-reviewed outlets and that described original research evaluating animals for detection of human diseases were included in the review. Most relevant studies have assessed dogs as detectors of various forms of cancer. Other researchers have targeted bacteriuria, Clostridium difficile, hypoglycemia, and tuberculosis. Nematodes and pouched rats were the only exceptions to canine detectors. Of the 28 studies meeting inclusion criteria, only 9 used operationally feasible procedures. The most common threat to operational viability was the use of a fixed number of positive samples in each sample run. Most reports included insufficient information for replication or adequate evaluation of the validity of the findings. Therefore, we have made recommendations regarding the type of information that should be included when describing research in this area. The results of this systematic review suggest that animal detectors hold promise for certain diagnostic applications but that additional research evaluating operationally viable systems for olfactory detection of human diseases is necessary.