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Bone is a unique biological composite material made up of a highly structured collagen mesh matrix and mineral deposits. Although mineral provides stiffness, collagen’s secondary organization provides a critical role in bone elasticity. Here, we performed polarimetric analysis of bone collagen fibers using second harmonic generation (SHG) imaging to evaluate lamella sheets and collagen fiber integrity in intact cranial bone. Our polarimetric data was fitted to a model accounting for diattenuation, polarization cross-talk, and birefringence. We compared our data to the fitted model and found no significant difference between our polarimetric observation and the representation of these scattering properties up to 70µm deep. We also observed a loss of resolution as we imaged up to 70µm deep into bone but a conservation of polarimetric response. Polarimetric SHG allows for the discrimination of collagen lamellar sheet structures in intact bone. Our work could allow for label-free identification of disease states and monitor the efficacy of therapies for bone disorders.

 

More than 54 million Americans have or are at high risk of developing a metabolic bone disease; disorders of bone strength that leave individuals with fragile bones and disabilities. The gold standard to evaluate these diseases is dual energy x-ray absorptiometry, but this only measures mineral content. These diseases, however, impact collagen and mineral integrity which impede the bone’s ability to store hormones, proteoglycans, and glycoproteins imperative to homeostasis. We have established a second harmonic generation (SHG) polarimetric assay that describes bone collagen organization. To further our analysis, we propose multimodal optical evaluation of bone quality with third harmonic generation (THG) to measure osteocyte dendritic processes. This method of analysis could be used to evaluate the disease state of bone and response to therapy.