Birefringence of Articular Cartilage and the Distribution of Collagen Fibril Orientations

Abstract

Birefringence of the deep zone of pig tibial plateau cartilage has been measured by polarized light microscopy using monochromatic light and a Sénarmont compensator. Thin (20 μm) sections of the tissue were imbibed with mixtures of physiological saline and glycerol to separate the effects of form and intrinsic birefringence. Measured birefringence was plotted against liquid refractive index and the experimental points defined a parabola whose minimum corresponded to a refractive index for collagen of 1.48. When the tissue was imbibed with glycerol, the birefringence measurements provided a good estimate of intrinsic birefringence. The orientation distribution function, g(ϕ), of collagen fibrils in this zone was then determined by X-ray diffraction. Thus X-ray diffraction yielded the form of g(ϕ) which corresponded to the measured intrinsic birefringence. Using this result the intrinsic birefringence of a hypothetical cartilage site with perfectly oriented collagen fibrils was calculated to be (6 ± 1) x 10^-4. Birefringence measurements on further tissue sites could then be converted into standard deviations of g(ϕ), assuming a Gaussian distribution.