Purpose: This study was performed to determine the accuracy of measurements of bone mineral density (BMD) and cortical bone thickness (CBT) at miniscrew implantation sites in small animals and to verify the usefulness of in vivo microcomputed tomography (micro-CT). Materials and Methods: Rat femurs were scanned before and after placing miniscrews using in vivo micro-CT. The images were superimposed using a subtraction method with bone volume measurement software. At each screw site, the total BMD was calculated as the average BMD of a cylinder 1.6 mm in diameter and 2.0 mm in depth, while the cortical BMD was the average BMD of a cylinder 1.6 mm in diameter and 1.0 mm in depth. CBT was measured three times on transaxial images of the rat femurs, and the average value was used. All miniscrews were placed using the maximum torque, verified with a digital torque tester. To verify the usefulness and accuracy of in vivo micro-CT, CBT on micro-CT images was compared with that measured on histologic sections. Results: Significant correlations were observed between placement torque and cortical BMD (R = 0.572), total BMD (R = 0.732), and CBT (R = 0.788). There was a significant correlation between CBT measured via field-emission scanning electron microscopy images and CBT measured with in vivo micro-CT (R = 0.974). Conclusions: The BMD over a narrow range can be measured accurately in small animals with high reproducibility of the trabecular structure using a combination of high-resolution in vivo micro-CT and image superimposition using a three-dimensional subtraction method.