The purpose of this study was to evaluate the effects of boundary conditions and specimen preparation on human rib testing. This was done by performing 48 three-point bending tests on 24 matched human rib specimens. The rib specimens were dissected from anterior and lateral regions of ribs 8-10 of three male human thoraces. Both specimens of each matched set were then potted side by side in a polyvinyl chloride (PVC) square pot filled with PMMA fast cast compound while using a custom jig to ensure the matching orientation. Once the specimens were potted, a microCT was used to obtain a detailed cross-sectional image of each specimen at the point of the impactor blade contact. The cross-sectional image was then thresholded and a custom Matlab code was used to calculate the area moment of inertia and distance to the neutral axis. Next, the matched specimens were randomly divided into three test groups. In the first test group, matched specimens were tested to determine the effects of leaving the periosteum intact versus removing the periosteum. In the second test group, matched specimens were tested to determine the effects of placing a strain gage on the tension side of the specimen. In the third test group, matched specimens were tested to determine the effects of immersing the specimens in saline for five days versus wrapping the specimens in saline soaked gauze. The soft tissue and periosteum were not removed in the second test group. The specimens were tested using a servo-hydraulic material testing machine (MTS) and a three-point bending setup, in which the potted specimen end was pinned and the other end was simply supported. Specimens were oriented so that the specimen was bent inside-out by the impactor blade. The impactor displacement rate of 7 in/s yielded a strain rate of approximately 0.5 strain/s, which is similar to that seen in a belted 35 mph automotive crash. In the first test group, a paired t-test showed that there was no statistical difference in the area moment of inertia (p=0.60), distance to the neutral axis (p=0.29), peak moment (p=0.31), peak stress (p=0.42), or peak impactor displacement (p=0.14) between specimens with an intact periosteum versus no periosteum. In the second test group, a paired t-test showed that there was no statistical difference in the area moment of inertia (p=0.76), distance to the neutral axis (p=0.20), peak moment (p=0.68), peak stress (p=0.34), or peak impactor displacement (p=0.91) between specimens with a strain gage versus no strain gage. In the third test group, a paired t-test showed that there was no statistical difference in the area moment of inertia (p=0.35), distance to the neutral axis (p=0.42), peak moment (p=0.45), peak stress (p=0.98), or peak impactor displacement (p=0.75) between specimens immersed in saline for five days versus specimens wrapped in saline soaked gauze. In summary, this procedure proved to be a very accurate means of preparing matched rib specimens and showed that neither the presence of the periosteum, strain gage, nor the two hydration conditions have a significant influence on the biomechanical properties of human rib specimens.