Objective To explore the acute toxicity effect of CKLF1-C19, a polypeptide of chemokine-like factor 1 (CKLF1), on the KM mice. Methods A total of 40 KM mice, half male and half female, were randomly divided into 2 groups. The mice in the experimental group were injected with CKLF1-C19 at a dose of 25 mL/kg (100 μg/mL, 1 mg/mL and 10 mg/mL) through the tail vein, and those in the control group received an equal volume of sterile saline solution. Changes in the body weight of the mice were recorded the day after treatment, and the general conditions of mice in the experimental group were observed closely and compared with the normal group. Then blood samples were taken from the abdominal aorta to measure liver and kidney function. Tissue samples of liver, kidney, spleen and lung were taken for histopathological examination by HE staining. Results In the maximum tolerance test, the mice of the two groups were in good condition, and their body weight was increased gradually, without significant difference between the experimental group and the control group (P > 0.05). There was no death within 14 days. The blood biochemical indexes of liver and kidney function showed no significant differences between the two groups (P > 0.05). The gross appearances of heart, liver, kidney, spleen and lung were normal in the two mouse groups, and the pathological examination with HE staining showed normal clear structure with no obvious changes in these organs of each group. Conclusions Our results demonstrate that CKLF1-C19 has no acute toxicity effect on mice.