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Suitable indentation profile
Please request
the chapter “Mechanical testing of soft tissues by indentation” (
Nelda Antonovaite,
"Exploring the mechanical microenvironment of the brain by dynamic indentation
”) which gives the
background for selecting indentation profile.
5.3
Sample stability
Additional to the environment, the sample itself can also introduce a measurement error.
Sample temperature and integrity
When attempting to measure samples that come fresh out of the incubator please allow some time for
readjustment to the Pavone temperature. This applies even more so to larger temperature gradients: if
medium or samples come out of a refrigerator, let the sample temperature stabilize to room temperature
before initiating the measurement.
The sample itself can also have unstable mechanical properties. It might be swelling if just hydrated or if
there is a change in the osmolarity of the medium or if the sample is degrading. When measuring a highly
heterogeneous sample, such as decellularized ECM, porous scaffolds or 3D-printed structures, extra care
should be taken in selecting the appropriate cantilever stiffness, tip size and indentation settings. For
example, when working with samples that have a porous surface, selecting a tip diameter that is either
much smaller or larger than the average pore size avoids having the tip trapped in the sample.
Sample immobilization
Next to a stable sample, it is important to immobilize the sample to eliminate lateral and height drift
during and between indentations. Indenting drifting or floating samples results in unreliable indentation
measurements such as high noise or irregular curves. Especially when performing automated matrix scans
make sure the sample is immobilized correctly. For more information about how to mount the sample,
please read the separate document called “Sample preparation protocols”.
Adhesive samples
Some hydrogels and tissues are very sticky which will result in a large distance needed for “Z above surf”
in order to perform the measurements out of contact. Also, there might be some snap-on behavior in the
loading curve which makes the calculation of the contact-point less precise. To decrease the effect of
adhesion, you can use a stiffer probe as the attractive forces will have a smaller effect on the cantilever.
However, using stiffer probe results in less precision for the soft samples at low indentation depths.
Furthermore, adhesion can be minimized by cleaning the probe from the dirt, coating the sample with 5%
BSA or coating the tip with 1% pluronic solution, where both of them make it hydrophobic.
Furthermore, the Hertz model assumes that the sample is not adhesive, thus, if adhesion is large, please
use the JKR model available in the DataViewer as it can estimate the contact area better. For more
information about adhesion and coating, please read
the separate document called “Sample preparation
protocols”.
Finally, ‘live calibration’ function in the maintenance should be turned off when measuring adhesive
samples (see Section 6.2).
