Downhill treadmill running is a commonly used method to cause exercise-induced muscle damage, especially in rodents. Previous studies have evaluated which muscles in rats are more prone to damage. However research using downhill run mice (DHR) has shown some inconsistencies in which muscle is best analyzed for damage. Purpose: The purpose of this study was to quantify the damage in various muscles in a mouse after a single bout of DHR. Methods: Male FVB mice (5 months) were injected with Evans Blue dye (EBD) and then either used as control (CON) or run downhill (-16°) at 20 meters per minute (m/min) for 30 minutes. Twenty-four hours after exercise, the gastrocnemius, soleus, plantaris, tibialis anterior (TA), quadriceps, and triceps brachii muscles were harvested (n = 6 per group per muscle). Cross-sectional slices were obtained, fixed, and mounted to analyze EBD infiltration, dystrophin (Dys), and centralized nuclei. The samples were then imaged using a fluorescent microscope. The entire sample was captured using 20x magnification, and the total number of cells, EBD+, Dys-, and centralized nuclei, were counted. A blood sample was collected to measure plasma creatine kinase (CK) activity. Results: Total number of cells was not different between groups (p > 0.05). No significant difference in any of the markers of muscle damage was found in any muscle between CON and DHR (p > 0.05). Conclusion: These data suggest that DHR does not induce muscle damage in adult (5 months) male FVB mice.