Image-guided Raman spectroscopic recovery of canine cortical bone contrast in situ

Abstract

Raman scattering provides valuable biochemical and molecular markers for studying bone tissue composition with use in predicting fracture risk in osteoporosis. Raman tomography can image through a few centimeters of tissue but is limited by low spatial resolution. X-ray computed tomography (CT) imaging can provide high-resolution image-guidance of the Raman spectroscopic characterization, which enhances the quantitative recovery of the Raman signals, and this technique provides additional information to standard imaging methods. This hypothesis was tested in data measured from Teflon tissue phantoms and from a canine limb. Image-guided Raman spectroscopy (IG-RS) of the canine limb using CT images of the tissue to guide the recovery recovered a contrast of 145:1 between the cortical bone and background. Considerably less contrast was found without the CT image to guide recovery. This study presents the first known IG-RS results from tissue and indicates that intrinsically high contrasts (on the order of a hundred fold) are available.

Fig. 1

(a) Reflectance mode instrumentation for Raman measurements is shown using a ring/disk geometry. The ring diameters can be varied to sample multiple depths for tomographic reconstruction. (b) Transmission mode configuration is shown using a rectangular array of collection fibers at 180 degrees from the line source illumination. Multiple projections across the tissue can be obtained by rotating the sample with respect to the source-collection fiber setup.

Fig. 2

(a) Schematic representation of the ring/disk measurements on the canine limb is shown using surface rendering from CT images. The ring diameter was varied between 6 and 16 mm for 10 different separations between source ring and collection fibers. (b) Representation of the transmission measurements from the canine tibia is shown. A total of 13 projections were obtained at 8° intervals around the limb.

Fig. 3

Cross-section of the reconstructed volume showing IG-RS estimates of the tissue phantom imaged using the ring/disk probe. The Teflon® inclusion was recovered with a contrast of 460:1 with respect to the background agar/1% Intralipid® gel.

Fig. 4

Surface rendering of bone and surrounding tissues. The outer limb surface was used to generate a volumetric mesh for computation.

Fig. 5

(a) Bone scores computed from reflectance measurements. (b) Bone scores computed from transmission measurements. The score plots indicate the correlation between the spectra obtained at each collection fiber and the bone spectra. (c) The bone scores were calibrated before the reconstruction using a scaling factor obtained from the phantom data, which compensated for the misfit between the FEM model and the measured data.

Fig. 6

A cross-section of the reconstructed IG-RS estimates using (a) reflectance measurements from the ring/disk configuration and (b) transmission measurements using the rectangular array of collection fibers. Recovered contrast between the bone and background skin was more than 100-fold higher using transmission measurements than using reflectance data, as seen from the scale for reconstructed Raman yield.