2.6. Microstructural analysis

Following mechanical testing, the chord tissues were fixed in 10% neutral buffered formalin for at least 24 h and dehydrated in a series of ethanol solutions of varying concentrations. The intact portion of each sample was processed in paraffin, sectioned into two 5-micron thick samples in both the radial and longitudinal directions, and then embedded in paraffin wax. Hematoxylin and Eosin (H&E) staining was carried out to visualize tissue integrity and orientation of the samples. Verhoeff–Van Giesson (VVG) staining was performed to visualize the collagen and elastin structures. Digital images of each slide were obtained utilizing a Zeiss Axio Scope.A1 microscope coupled with a Zeiss AxioCAM RMm microscope camera. The crimped structure of the collagen fibers were also assessed from the H&E stained samples and observed with a Zeiss Observer.D1 inverted microscope.