This study explores relationships between 'in vivo ' speed of sound and 'in vitro' material properties in the equine third metacarpus. Speed of sound, ultimate tensile strength, elasticity and bone mineral density were measured for discrete sites in 25 bones. Experimental procedures were simplified through development and validation of a method for determining effective centroid locations in whole bone samples and a modular strain measurement clip. Although the speed of sound measurements were highly site dependent, statistically significant correlations were found with the variables in 4 of 9 experimental sites. Speed of sound correlated with bone mineral density (r2 = 0.25, p < 0.01), and elasticity (r2 = 0.14, p < 0.05) at the dorsal mid diaphysis. A strong correlation (r2 = 0.48, p < 0.001) was observed between speed of sound and ultimate tensile strength in the dorsal distal site. Results indicate quantitative ultrasound is sensitive to differences in equine third metacarpal material properties.