Quantification of the second-order nonlinear susceptibility of collagen I using a laser scanning microscope.

Characteristic changes in the organization of fibrillar col- lagen can potentially serve as an early diagnostic marker in various pathological processes. Tissue types containing collagen I can be probed by pulsed high-intensity laser radiation, thereby generating second harmonic light that provides information about the composi- tion and structure at a microscopic level. A technique was developed to determine the essential second harmonic generation SHG param- eters in a laser scanning microscope setup. A rat-tail tendon frozen section was rotated in the xy-plane with the pulsed laser light propa- gating along the z-axis. By analyzing the generated second harmonic light in the forward direction with parallel and crossed polarizer rela- tive to the polarization of the excitation laser beam, the second-order nonlinear optical susceptibilities of the collagen fiber were deter- mined. Systematic variations in SHG response between ordered and less ordered structures were recorded and evaluated. A 500m-thick z-cut lithiumniobate LiNbO3 was used as reference. The method was applied on frozen sections of malignant melanoma and normal skin tissue. Significant differences were found in the values of d22, indicating that this parameter has a potential role in differentiating between normal and pathological processes. © 2007 Society of Photo-Optical