Monkeys are frequently used in experimental transplantation research because of their physical traits and availability. As ABO incompatibility may result in humoral injury, it is important to identify the ABO blood typing of monkeys before transplantation. However, monkeys lack expression of ABH antigens on red blood cells, which makes accurate determination of the blood type difficult. The gel agglutination assay has been widely used as a routine blood grouping test clinically for more than 10 years. In this study, we evaluated the efficacy and the interference factors of using the gel system (including the direct gel system and the reverse gel system) for ABO typing in rhesus monkeys (n = 38) and cynomolgus monkeys (n = 26). Immunohistochemistry assay was used to obtain the accurate blood type data of monkeys. The results revealed that the direct gel system was ineffective in blood typing of monkeys, whereas the reverse gel system assay, which is based on preabsorbed serum, provided reproducible results that were confirmed by histologic analysis. We conclude that the reverse gel system assay with use of preabsorbed serum is a simple and reliable method for ABO typing of monkeys.