Myopia (also called near-sightedness) is an irreversible ocular abnormality that occurs when the eye is excessively elongated, resulting in defocused images and vision. When myopia progresses to pathological myopia, the possibility of permanent vision loss is extremely high. Therefore, it is important to identify early myopic eyes likely to progress to pathologic myopia and vision loss to start treatment and intervene as soon as possible. As myopia progresses, posterior eye elongation is caused by changes in the microstructural properties of the sclera including decreased collagen density and fibril diameters. High-frequency quantitative ultrasound (QUS) allows measuring biomechanical parameters associated with changes in tissue microstructure. This study investigated microstructural changes occurring in the posterior sclera in myopic guinea pig eyes using two independent QUS approaches at 80-MHz: envelope statistics quantification using the Homodyned-K distribution and backscatter coefficient quantification using a cylindrical Gaussian form factor. QUS parameters suggest changes in collagen fiber size occur in myopic sclera and is associated with microstructural changes in the scleral collagen fiber homogeneity and density. In conclusion, high-frequency QUS has the potential to quantitatively characterize microstructural changes that occur in the posterior sclera during myopia development and/or progression.