Exposure to methylmercury (MeHg) can result in detrimental health effects in wildlife. With advances in ecological indicators and analytical techniques for measurement of MeHg in a variety of tissues, numerous processes have been identified that can influence MeHg concentrations in wildlife. This review presents a synthesis of theoretical principals and applied information for measuring MeHg exposure and interpreting MeHg concentrations in wildlife. Mercury concentrations in wildlife are the net result of ecological processes influencing dietary exposure combined with physiological processes that regulate assimilation, transformation, and elimination. Therefore, consideration of both physiological and ecological processes should be integrated when formulating biomonitoring strategies. Ecological indicators, particularly stable isotopes of carbon, nitrogen, and sulfur, compound-specific stable isotopes, and fatty acids, can be effective tools to evaluate dietary MeHg exposure. Animal species differ in their physiological capacity for MeHg elimination, and animal tissues can be inert or physiologically active, act as sites of storage, transformation, or excretion of MeHg, and vary in the timing of MeHg exposure they represent. Biological influences such as age, sex, maternal transfer, and growth or fasting are also relevant for interpretation of tissue MeHg concentrations. Wildlife tissues that represent current or near-term bioaccumulation and in which MeHg is the predominant mercury species (such as blood and eggs) are most effective for biomonitoring ecosystems and understanding landscape drivers of MeHg exposure. Further research is suggested to critically evaluate the use of keratinized external tissues to measure MeHg bioaccumulation, particularly for less-well studied wildlife such as reptiles and terrestrial mammals. Suggested methods are provided to effectively use wildlife for quantifying patterns and drivers of MeHg bioaccumulation over time and space, as well as for assessing the potential risk and toxicological effects of MeHg on wildlife.