Collagen fiber spatial orientation mapping using polarization-sensitive SHG microscopy

摘要

Collagen, the main structural protein in vertebrates, possesses different structural organization that is responsible for specific functions of the tissues. Polarization dependence of the second harmonic generation (SHG) signal on spatial orientation of optically nonlinear materials, such as collagen, provides information on characteristic organization and architecture not available from intensity measurements alone. Here we describe a simple approach for determining both the azimuth and elevation angles of collagen fiber orientation in biological tissues. Azimuth angle of the fiber orientation is determined as an orthogonal angle to the laser polarization direction, when laser-induced total SHG signal is minimal, whereas the elevation angle is estimated from the ratio of the minimal SHG intensity to the intensity when laser polarization and fiber directions are parallel to each other. Using this approach pixel-resolved mapping of the spatial orientation of collagen fibers in tendon and cornea is demonstrated. The new approach may be used for analyzing of biological tissues in vivo. Spatial orientation mapping method provides additional information concerning fiber organization, and may be incorporated into nonlinear optical imaging systems.