Biomedical Engineering Reference
In-Depth Information
Appendix A
Accurate, reproducible and quantitative image acquisition with an AFM requires some
care. The most important procedure to enable reproducible and accurate results is cali-
bration of the AFM. In order to carry out such procedures, accurate standards and
references are required. Standards are used to ensure that the absolute measurements are
correct, while references ensure that the instrument is giving consistent results. Most AFM
calibration procedures involve using the AFM to make a measurement on a well-known
sample such as the standards described in this chapter. Scanner calibration and certifica-
tion procedures are described in Appendix B. Different standards for AFM calibration are
available for
X-Y
and
Z
axis calibration. Apart from calibration of the scanner, the most
important calibration is measurement of the tip of the probe used. The probe in AFM
experiments is manufactured with rather a large tolerance, so reproducibility between
probes can be quite low. However, the sharpness and aspect ratio of the probe used is one
of the factors that can define the achievable resolution in an AFM experiment. Standard
samples described in this chapter can enable measurement of probe radius, which greatly
helps in understanding and controlling image quality. The samples and products described
in this chapter are not the only ones that can be used for calibration, any sample with
reproducible and well-known dimensions may be used for AFM characterization. Since
any printed list of internet addresses is likely to rapidly go out of date, there is updated
version of the calibration specimen supplier listing on the website accompanying this topic
A1 Z axis calibration
Calibration standards and references are needed to calibrate AFMs in the vertical axis. For
calibrations greater than 10 nm step height, standards or references are typically fabricated
by lithographically etching patterns in a silicon or quartz substrate. It is also possible to
find silicon dioxide coated with a uniform layer of metal. When calibrating the instrument
for
Z
height measurements below 10 nm, nanoparticles with low polydispersity or atomic
terraces of silicon or HOPG may be used as a reference specimen. Figure A1 shows an
AFM image and measurement of a
Z
axis calibration specimen consisting of a pit of
ca
.
16 nm depth. Table A1 lists some
Z
axis calibration specimens that are commercially
available.
