NANOINDENTATION OF BONE - Nanoindentation with Biological Applications - page 221

4.3. Modulus variation with mineral composition
The elastic modulus positively, and non-linearly, correlates with the
mineral content in bulk samples of bone. 132,133 Nanoindentation provides
a unique approach to study this same relationship at the tissue-level,
whereby measurements of mineral volume fraction or composition can
be collected at a similar spatial, micrometer-scale resolution.
Correlation of mineralization with nanomechanical properties can
be achieved through combined analysis using nanoindentation and
qBSE imaging. 50,172 The mean grey level in the effective contact area for
each indent site within a grid array, mapped over a qBSE image of
the indented region ( Fig. 7-6a ) , yields the mineral volume fraction
through the use of calibrated standard materials. 50,108,127,173 The qBSE
image brightness is dependent on the mean atomic number at a given
point, which is representative of mineral content, where more highly
mineralized regions appear brighter than more poorly mineralized
regions.
Correlation with nanoindentation modulus is achieved by collecting
qBSE information in areas that match the effective contact area for each
indent site. 50,173 Additional refinements to the analysis are provided by
segregating data into populations by the type of tissue on which the
indents lay. The example provided here shows data for two grid arrays
that are separated, for each matched E - mineral V f data point, into
immediate subchondral bone (B) and articular calcified cartilage (C)
( Fig. 7-6b ). Additional classifications, such as by tissue type or extant
versus newly formed bone (through the use of florescent labels), may
improve the fidelity of the analysis. As previously mentioned in Section
3.2, a grid indentation approach provides a statistical sampling of a
composite material, this method of analysis lacks bias and serves as an
effective tool to randomly sample within a region of interest. 126 Plots of
E versus mineral V f thus reveal information about the relationship
between nanomechanical properties and mineralization ( Figs. 7-4 and
7-6 ) .