Objective To compare the hematological parameters, biochemical profiles, histopathological characteristics, and cytokines associated with lipid metabolism in 8-weeks-old specific pathogen-free (SPF) grade and germ-free(GF) golden hamsters. Methods Twenty 8-weeks-old SPF grade and GF golden hamsters, with equal numbers of males and females, were utilized in this study. Serum cytokines associated with routine blood parameters, blood biochemistry, and lipid metabolism were quantified using automated hematology and biochemical analyzers, and enzyme-linked immunosorbent assay kits. The cecum, small intestine, lung, spleen, forestomach, and glandular stomach were examined by histopathology. Results We compared the result between SPF grade and GF golden hamsters at 8 weeks of age. Regarding hematological parameters, females showed significant differences (P<0.05) in white blood cell count, hemoglobin concentration, mean corpuscular hemoglobin (MCH), and mean platelet volume between the two groups, and highly significant differences (P<0.001) in MCH concentration (MCHC), platelet distribution width (PDW), and lymphocyte count (LYM). Males showed significant differences (P<0.05) in mean corpuscular volume, MCH, LYM, neutrophil count, basophil count, and basophil percentage, and highly significant differences (P<0.001) in MCHC, PDW, lymphocyte percentage, and neutrophil percentage. In terms of biochemical parameters, females showed significant differences (P<0.05) in low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol, fasting insulin (FINS), and glycosylated serum protein (GSP), and highly significant differences (P<0.001) in triglycerides (TG). Males showed significant differences (P<0.05) in GSP levels and highly significant differences (P<0.001) in total cholesterol, TG, LDL-C, FINS, and C-peptide levels. For serum cytokines related to lipid metabolism, females showed significant differences (P<0.05) in interleukin (IL)-10, adiponectin (ADP), and IL-6 levels, and highly significant differences (P<0.001) in high sensitivity C-reactive protein (hs-CRP). Males showed significant differences (P<0.05) in hs-CRP levels and highly significant differences (P<0.001) in ADP and tumor necrosis factor-alpha levels. Hematoxylin/eosin staining showed that the cecal muscle layer was thinner and the number of crypts attached to the mucous membrane was reduced in GF compared with SPF grade golden hamsters. In addition, the ileocele was enlarged and the number of goblet cells was increased, the alveolar septum was widened, immune cells in the white pulp of the spleen were increased, and the blood content in the blood sinuses was increased. There was also thinning of the anterior gastric mucosa and the basophilic strength of the glandular gastric tube gland was weakened. Conclusions This study established the differences in routine blood parameters, blood biochemistry indicators, histopathological features, and cytokines associated with lipid metabolism between 8-weeks-old SPF grade and GF golden hamsters, to establish preliminary reference ranges.