Evaluating amino acid requirements, specifically threonine requirements, in horses will enable better feed formulation and result in economic production, improved animal health, and reduced environmental pollution. However, the current knowledge of protein and amino acid requirements in horses is still limited. Because horses have a unique digestive system and consume a variety of feed ingredients, their protein digestibility may be affected than other species by different feed composition, and thus amino acid requirements are susceptible to vary between situations. Therefore, a careful evaluation of amino acid requirements with a proper method is needed for various conditions. This review will also provide comprehensive information that needs to be considered when designing an amino acid requirement study in horses.