INSTRUMENTATION AND EXPERIMENTATION - Nanoindentation with Biological Applications - page 72

over the experiment. Thus, pixellated maps of mechanical properties
inferred from AFM-enabled indentation should be considered qualitative
estimates of mechanical variation that, unless carefully calibrated, to not
reflect the mechanical properties of pixel-sized biomaterial volumes.

3.5 . Sample mounting and preparation

As in the case of instrumented indentation (Section 2.5 of this chapter),
biomaterial sample preparation is a critical part of accurate mechanical
characterization. Only issues different from, or in addition to, those
already presented for instrumented indentation will be considered here.
The change in fluid levels or salinity will affect the positional
stability of the AFM cantilever. It is important to maintain the sample
and the entire fluid cell under a constant volume of fluid, either by
adding fluid periodically upon evaporation or, preferably, by adopting a
closed cell configuration that minimizes evaporation.
In certain experiments, one may require exchange of fluids during
mechanical analysis. To minimize mechanical agitation, injection via
manually depressed syringe needles is preferable to mechanical pumps.
To secure biomaterial samples to rigid, flat surfaces required of
AFM-enabled indentation, several options can be considered. In general,
the goal is to maximize the adhesion strength of the sample to the rigid
surface without denaturing or otherwise structurally altering the
biomaterial. Proteinaceous samples can be secured with CellTak™ or
similar biological adhesives, and can sometimes be physically adsorbed
to surfaces via mild centrifugation or even drying and rehydration. As a
rule, the less mechanical and physical stress exerted on the sample, the
more likely that the surface characterized via AFM-enabled indentation
will accurately reflect the properties of the intact biomaterial sample.

4. Summary and Conclusions

This chapter has considered experimental issues relevant to contact
mechanics testing techniques for biological materials. Separate sections
have examined instrumented indentation (Section 2) and AFM-based
testing (Section 3). Emphasis has been on issues of load and