Greater understanding of tympanic membrane (TM) biomechanics has the potential to guide future advances in medical
technology related to its surgical repair (myringoplasty). The pars tensa of the TM is a composite structure with two
collagen fiber layers that provide the main scaffolding for the TM. The external layer is arranged in an approximately
radial configuration, and the other is arranged in an approximately circumferential configuration. A more detailed
knowledge of collagen fiber orientation and volume fraction could greatly improve existing mechanical simulations of
the TM. To address this, we employed multiphoton microscopy (MPM) imaging of the TM in two modalities: second
harmonic generation (SHG) and two-photon fluorescence (TPF). The unique spectral signature of SHG allows selective
imaging of collagen fibers. TPF also produces images of fibrillar-type collagen but lacks the specificity of SHG. Both
the SHG and TPF images show patterns of collagen organization in the TM that match expected results with respect to
both orientation and size. Through MPM, we intend to accurately determine the collagen fiber layer thickness, density,
and orientation as a function of radial position and quadrant location.
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