Collagen is the primary structural component of musculoskeletal tissues such as tendons and cartilage. It exists in a hierarchical structure, and its base unit, the collagen molecule, can become damaged or denatured in various musculoskeletal injuries and diseases. Therefore, being able to quantify collagen denaturation is essential for understanding the etiology and progression of musculoskeletal ailments. A technology for easily characterizing denatured collagen is collagen hybridizing peptides (CHPs), which bind selectively with denatured, but not intact collagen. This dissertation focused on applying CHPs to characterize denatured collagen during tension throughout the collagen hierarchy in tendons.

Aim 1 developed a rapid, high-throughput method of quantifying denatured collagen using CHPs. We created a microplate assay that maintained the accuracy of the current gold-standard technique for quantifying denatured collagen, while reducing the process from 3+ days to less than 24 hours. Furthermore, we demonstrated that this method can be utilized in various collagenous tissues with different collagen types. The outcomes of Aim 1 were used in Aim 2 to identify the onset of collagen denaturation during tensile load relative to the stress-strain curve in two different types of tendons. Tendons are divided into two categories with unique biomechanical and biochemical properties – positional and energy-storing. We found that despite these differences, the onset of collagen denaturation occurred at the yield point in both tendon types during monotonic stretch. This indicated that the failure mechanism is the same in both positional and energy-storing tendons. We were interested in whether this trend is maintained throughout the collagen hierarchy, so in Aim 3, we performed tensile tests on collagen fibrils using a microelectromechanical system (MEMS) device. Collagen fibrils are one level above collagen molecules in the collagen hierarchy, allowing us to more directly probe the effect of tension on collagen denaturation. We found that collagen fibrils from both tendon types exhibited increased amounts of denatured collagen only when stretched beyond the yield point of the stress-strain curve, similar to tissue level results. The results of this dissertation will help researchers understand how musculoskeletal injuries and diseases occur and progress, leading to improved clinical outcomes in the future.