Objective To explore differences between three- and five-classification automatic hematology analyzers and manual counting to measure physiological indexes of peripheral blood erythrocytes of tree shrews and establish a reliable reference. Methods The normal reference value range of the number of red blood cells, mean corpuscular volume, hematocrit, hemoglobin concentration, mean corpuscular hemoglobin concentration, and mean corpuscular hemoglobin in the peripheral blood of 20 healthy adult tree shrews were measured by the manual method , five-classification automatic blood cell analyzer BC-6800, and three-classification automatic blood cell analyzer UINT-3010. The consistency of the two method was judged by a Bland-Altman diagram and Passing-Bablok regression analysis. Results Significant differences in the number of red blood cells, mean corpuscular volume, hemoglobin concentration, and mean corpuscular hemoglobin concentration were observed between the manual method and five-classification automatic hematology analyzer (P < 0. 001). No systematic difference was found between the two method , but there was a proportion difference. No statistical difference in hematocrit or mean corpuscular hemoglobin was found, and the two method had good consistency, no systematic difference, and a proportion difference. Significant differences were observed between the manual method and three- classification automatic hematology analyzer except for the number of red blood cells (P < 0. 001). Conclusions The normal reference range of red blood cell-related physiological indexes measured by hand provides a reference to study the tree shrew. The BC-6800 automatic blood analyzer is more accurate and reliable than the UNIT-3010 to measure red blood cell-related indicators, which can be used for rapid and effective analysis of a large number of samples, but it cannot completely replace the manual method. Only the combination of the two can ensure the accuracy and reliability of test result.