Zinc Oxide (ZnO) nanorods have been prepared and proposed for collagen Type I reinforcement. The insertion of ZnO has been confirmed by an x-ray Diffraction pattern. A nano-indentation technique implemented through an Atomic Force Microscopy has been effectively handled to probe a mechanical behavior of soft tissue like collagen. Measurements at different temperatures for collagen thin films filled with different amounts of ZnO reveals that ZnO significantly develops Young’s modulus from 9 GPa to 18 GPa. Without ZnO, the temperature increment leads to the dehydration of pure collagen while the reinforced ones were still elastic for cell growth. ZnO-reinforced collagen has, therefore, been improved both inherent rigidity and its stability against ambient temperature. The finding in this work suggests that nanoscale ZnO is good candidate for collagen reinforcement which is crucial for tissue engineering technology