Objective Differences in peripheral blood and immune cell phenotype were compared between aged and young Sprague-Dawley rats. Methods The peripheral blood cell count, leukocyte classification, immune organ coefficient, immune cell phenotype in peripheral blood and spleen, and p16 expression in spleen T cells were detected in 8-week-and 12-month-old rats. Results Compared with young rats, the peripheral blood leukocyte count and percentage of lymphocytes in aged rats were decreased, and the percentages of erythrocytes, hemoglobin, neutrophils, eosinophils, and basophils were increased. However, there was no difference in the percentages of platelets and monocytes between groups. Additionally, there were no differences in thymus coefficient and spleen coefficient between young and old rats. Peripheral blood immunophenotyping indicated helper T cells, regulatory T cells, natural killer cells, and monocytes were increased, whereas B cells were decreased in aged rats compared with young rats. However, there was no significant difference in cytotoxic T cells. In old rats, markedly lower numbers of B cells (CD45RA) were present in the spleen compared with young rats, and there was no significant difference in CD3 cells between the groups. In addition, the expression of P16mRNA in spleen T cells of aged male rats was significantly higher than in young male rats. Conclusions In aged rats, the differentiation of peripheral blood cells was deviated, the proportion of neutrophils was increased, and the proportion of lymphocytes was decreased compared with young rats. The proportion of regulatory T cells in the peripheral blood was increased, and the proportion of B lymphocytes was decreased in the peripheral blood and spleen in aged rats compared with young rats. These changes in innate immunity and acquired immunophenotype suggest that the immune function of aged rats was decreased and the regulation of immune function was disordered, similar to changes in the immune system of elderly population. The expression of P16 in spleen T cells was increased in aged rats, and therefore might be used as a biomarker for immune senescence. This study provides primary data for the study of age-related diseases and ageing animal models.