Bone quality is an important measure of welfare in laying hens for researchers, veterinarians, and commercial producers who bear the responsibility of providing them with care. Breeding practices that have resulted in high egg productivity of laying hen strains have resulted in an increased susceptibility to developing osteoporosis. Osteoporosis, a decrease in mineralized bone, is accompanied by increased bone fragility that can lead to potentially painful bone fractures that can inhibit some aspects of the birds natural living, respiration and flight being two examples. Current methods of examining laying hen bone quality involve the implementation of diagnostic imaging and biomechanical assessments, most commonly conventional/digital radiography, computed tomography (CT), and the 3-point bending test. Current studies utilizing radiographic examinations to assess poultry bone health have largely relied on the use of scoring systems that are unique to the respective lab group conducting the research. This is apparent from reviews of the literature that have highlighted the discordance amongst reported scoring systems that use the presence or absence of fractures, fracture gaps, callus formation, bone deviations, and abnormal angulations of bone morphology to assign a qualitative score to a bone to characterize its damage status. The result of utilizing these varying scoring systems is that there is a lack of generalizability of the results and a subsequent inability to make comparisons between studies that would be beneficial in making efforts towards addressing the welfare concern. Additionally, scores do not allow the use of stronger statistical tests that can be performed with quantitative data based on discrete and continuous measures. Computed tomographic (CT) and biomechanical analyses are established, quantitative methods for characterizing the bone quality of laying hens. Reported analyses have assessed the 3 bone mineral density (BMD) of the keel, tibiotarsus, femur, and humerus; longitudinal morphology changes of the keel; cross sectional area of the keel and tibiotarsus; cortical thickness; and the angulation of the ventral margin of the keel using image reformatting. Current published methods of the assessment of keel and tibiotarsal bone quality include variations of assessing the BMD of different bone types. Total bone; the midpoint of the tibiotarsal diaphysis; and defined proximal, middle, and distal locations of the keel and tibiotarsus served as points for CT assessment of BMD/Bone mineral content (BMC) in the published literature. Less frequently however is CT muscle quality evaluated despite its potential to be correlated with bone health as is reported in human literature. Additionally, the 3-point bending test has been used as the primary method of assessing bone quality of laying hens biomechanically with differing testing parameters (fulcrum width and crosshead speed) used between studies. Given the knowledge gaps present in the literature pertaining to radiographic and CT assessments of poultry bone quality, the objective of the retrospective study detailed in Chapter II was to develop and assess the intra-observer repeatability of a standardized radiographic protocol for quantifying three types of KBD (keel bone fractures, ventral margin deviation, and dorsal margin angulation) using Horos open-source image analysis software and discrete and continuous variables. Objectives of the prospective study detailed in Chapter III were the following: 1) develop standardized protocols for quantifying keel bone, tibiotarsal, and muscle quality using CT and open-source image analysis software (Horos, https://horosproject.org); 2) test associations between CT cross sectional area of the muscle group surrounding the tibiotarsus and tibiotarsal BMD; 3) test associations between CT cross sectional area of the muscle group surrounding the keel and keel BMD; 4) develop a standardized protocol for quantifying biomechanical properties of tibiotarsal bones; and 5) test associations between biomechanical 4 measures and CT measures of bone quality. Our findings indicated that the radiographic protocol was repeatable and had the potential the increase the generalizability of results if applied to other studies. Implementation of the developed radiographic protocol that utilizes quantitative, discrete and continuous measurement variables would also allow for the use of stronger statistical tests. It was also found that associations exist between CT measures of tibiotarsal bone and muscle quality at the proximal location of the bone. Computed tomographic measures of bone and muscle quality were also predictive of biomechanical measures of bone quality of the tibiotarsus. These findings justify the use of the measure of the transverse, cross sectional muscle area of the tibiotarsus in future studies. This measure is not currently reported in the poultry literature, although current literature suggests there is a relationship between measures of bone quality and body weight. This measure, if utilized in studies aiming to address welfare concerns, has the possibility of identifying the effects of interventions on tibiotarsal musculature and can indirectly provide a measure of bone quality, further reducing the need for destructive testing. Additionally, findings further justify the exploration of the suitability of laying hens for use as translational research models for osteoporosis in humans.