Biomedical Engineering Reference
In-Depth Information
Chapter 4
Like all techniques, AFM requires some skill and practice to operate well, but learning to
measure an AFM image is quite easy, and usually just takes a few hours of instruction and
practice. Preparing the samples, setting up the instrument and scanning two to three
images can take only half an hour. However, if it is an unknown sample that was never
scanned by AFM before, it can take substantially more time to acquire useful images.
In this chapter we discuss the procedures that can make measuring AFM images easier.
This section does not replace the AFM manufacturer's user manual. Details specific to
each instrument can be found in those documents. Instead, here we show the overall steps
required for scanning a range of common samples, under typical conditions, and how to
optimize conditions to get the best images. This chapter covers the most common imaging
procedures; it focuses on contact mode and intermittent contact-mode AFM (IC-AFM).
Non-contact-mode AFM is currently used much less widely than IC-AFM, so it is not
explicitly covered here, but the imaging procedure is quite similar to that of IC-AFM. In
addition to imaging procedures, some details on obtaining force-distance curves are
included, as many users will also measure these. Figure 4.1 shows the major steps involved
in measuring an image in an optical lever-based AFM.
In general, sample preparation for AFM is very simple. For example, there is no need for
the sample to be coated, electrically grounded, stained, or to be transparent, as required for
some electron microscopic techniques. Some samples, such as thin films, can require no
sample preparation at all. Other samples, such as human cells, or very small nanoparticles,
may require considerable care in preparation for the best results. The 'rules' for prepar-
ation of samples for contact-mode AFM can be summarized as follows:
• The sample must be fixed to a surface. AFM is a surface technique, so all samples
require some kind of substrate. Some common substrates for AFM are discussed
below. If the sample consists of, or includes loose particles, these must be adhered to
the surface before scanning. If some material on the surface is not well fixed down, it
can lead to the AFM tip moving the material around on the sample surface. This can
lead to a 'sweeping' of the surface, eventually clearing the substrate, with the particles
being moved to the edge of the scan range. This sort of behaviour is particularly
common in contact-mode AFM, as the tip never leaves the surface, and it can apply
considerable lateral forces to the surface. Even if the sample is not 'swept' in this way,
moving material on the surface will lead to inconsistent images, and 'streaking' as the
tip encounters particles that are loose on the surface. It is also common for such
particles to be transferred from the surface to the tip under these conditions. This will
