We report the use of spatial arrays of single quantum dots (QD) as fluorescent probes to quantify deformations and
displacements of bone tissue components (e.g. collagen and carbonated apatite) at the nanometer to micrometer level
under mechanical load. Quantum dot bright emission and robustness allow nanometer localization and motion tracking
by center of gravity (COG) analysis. Coupons of milled cortical bone are loaded in a purpose-built dynamic mechanical
loading system that fits on a microscope stage. We used QD streptavidin conjugates to label the bone specimen prior to
mechanical loading. COG of the laser-induced QD fluorescence diffraction spot is measured and tracked in real time
(<0.1 sec) as the tissue is loaded quasi-statically. The technique has been validated by comparing the average values of
tangent elastic moduli obtained by the QD/COG method to measurements made with an attached micro-strain gage and
a calibrated load cell. Two or more colors of QD can be used to measure relative motions of different bone tissue
components, as well as to measure small out-of-plane motions that cannot be detected otherwise.
Background fluorescence can often complicate the use of Raman microspectroscopy in the study of musculoskeletal tissues. Such fluorescence interferences are undesirable as the Raman spectra of matrix and mineral phases can be used to differentiate between normal and pathological or microdamaged bone. Photobleaching with the excitation laser provides a non-invasive method for reducing background fluorescence, enabling 532 nm Raman hyperspectral imaging
of bone tissue. The signal acquisition time for a 400 point Raman line image is reduced to 1-4 seconds using electronmultiplying
CCD (EMCCD) detector, enabling acquisition of Raman images in less than 10 minutes. Rapid photobleaching depends upon multiple scattering effects in the tissue specimen and is applicable to some, but not all experimental situations.