Scanning Probe Microscopy (SPM)

John W. Cross

[ Applications ]

[ Choice of Technique ]

[ Accuracy and Calibration ]

[ Instrument Makers ]

[ Footnotes, References and Links ]

[ Books about SPM ]

[ Graphic equivalent ]


Scanning probe microscopy covers several related technologies for imaging and measuring surfaces on a fine scale, down to the level of molecules and groups of atoms.  At the other end of the scale, a scan may cover a distance of over 100 micrometers in the x and y directions and 4 micrometers in the z direction.  This is an enormous range.  It can truly be said that the development of this technology is a major achievement, for it is having profound effects on many areas of science and engineering.

These technologies share the concept of scanning an extremely sharp tip (3-50 nm radius of curvature) across the object surface. The tip is mounted on a flexible cantilever, allowing the tip to follow the surface profile.

As the tip passes over bumps in the surface, the vertical deflection of the cantilever (delta-z) is measured.

When the tip moves in proximity to the investigated object, forces of interaction between the tip and the surface influence the movement of the cantilever or are detected by selective sensors. Various interactions can be studied depending on the probe sensors used.

The three most common types of scanning probe techniques are:

Atomic Force Microscopy (AFM) measures the interaction force between the tip and surface. The tip may be dragged across the surface, or may vibrate as it moves.

Scanning Tunneling Microscopy (STM) measures a weak electrical current flowing between tip and sample.

Near-Field Scanning Optical Microscopy (NSOM) scans a very small light source very close to the sample.  Detection of this light energy forms the image.  NSOM can provide resolution below that of the conventional light microscope.

There are numerous variations on these techniques. AFM may operate in several modes which differ according to the force between the tip and surface:

Mode of Operation                Force of Interaction

contact mode                        strong (repulsive) - constant force or constant distance
non-contact mode                 weak (attractive) - vibrating probe
intermittent contact mode       phase or amplitude of cantilever oscillation is affected
friction mode                         frictional forces exert a torque on the scanning cantilever
magnetic force                       the magnetic field of the surface is imaged
thermal scanning                    the distribution of thermal conductivity is imaged 

In contact mode, the tip is usually maintained at a constant force by moving the cantilever up and down as it scans.  In non-contact mode or intermittent contact mode (tapping modeTM) the tip is driven up and down by an oscillator.  In non-contact mode, the bottom-most point of each probe cycle is in the attractive region of the force-distance curve.  In intermittent contact mode the bottom-most point is in the repulsive region.  Variations in the measured oscillation amplitude and phase in relation to the driver frequency are indicators of the surface-probe interaction.

To image frictional force, the probe is dragged along the surface, resulting in a torque on the cantilever.  To image the magnetic field of the surface, a magnetically-susceptible probe is used.  In other variations, the electric charge distribution on the surface or the surface capacitance is imaged.
For thermal scanning microscopy (TSM) the thermal conductivity of the surface with is probed with a bimetallic tip out of contact with the sample surface.

Several types of probes with different tips are used in scanning probe microscopy.  Tip selection depends on the mode of operation and on the type of sample.


These techniques have the ability to operate on a scale from microns down to nanometers and can image clusters of individual atoms and molecules. STM relies on the electrical conductivity of the sample, so at least some features on the sample surface must be electrically conductive to some degree. AFM is used for studies of non-conductors and is the technique more commonly used for studies of macromolecules and biological specimens. AFM has been used for measurements on a wide variety of sample types, including:

Inorganic and Synthetic Materials 
[ Natural surface topography ]
[ Surface Chemistry ]
[ Silicon wafers ]
[ Recording media ]

[ Buckyballs and Nanotubes ]
[ Crystalline Polymers ]
[ Diffraction gratings ]
[ Integrated circuits
Biological Materials
 Polymers and Polymer Matrix
[ Resins and gums ]
[ Muscle proteins ]
[ DNA ]
[ Plant  cell walls ]

Biological Structures
[ Bacterial flagellae ]
[ Chromatin and Chromosomes ]
[ Cell and membrane surfaces


Choice of Technique

The analytic mode is chosen based on the surface characteristics of interest and on the hardness or stickiness of the sample.  Contact mode is most useful for hard surfaces.  However, a tip in contact with a surface is subject to contamination from removable material on the surface.  Excessive force in contact mode can also damage the surface or erode the sharpness of the probe tip.  Near-contact mode has less tendency to deform a soft surface, but is more sensitive to environmental vibrations and to variations in the film of moisture that coats samples in a normal atmosphere.  Moisture or other thin liquid films expert an attractive capillary force on the probe as it is withdrawn from the surface.  In non-contact mode this force becomes significant.

Vibrating mode or intermittent contact modes are particularly suited for imaging soft biological specimen. However, biological samples are successfully imaged in the "harder" contact mode. Unfixed soft specimens are deformed in the z-dimension to a degree dependent on the imposed probe force, although spreading in the x-y plane may not be significant. Biological samples may be hardened to reduce probe-induced deformation by aldehyde fixation or frozen in a cryo-AFM.  See the Footnotes for further information on technique and sample preparation.

Accuracy and Calibration

Instrumental Factors. The performance of a scanning probe instrument is limited by a number of factors. A major factor is the resolution of the mechanical components used to move the tip and measure its position. The sharpness and stability of the probe tip determine the area of contact and the reproducibility of imaging. Obviously, environmental vibrations must be controlled to a high degree. In addition, most positioners depend on piezoelectric drive, which is subject to problems of non-linearity and to overshoot during rapid movements. The major manufacturers of SPM equipment have made substantial improvements in mechanical and electronic design. These improvements and advanced electronic calibration routines result in measurements that are more linear and accurate than the early models.   Mark VanLandingham (University of Delaware) has published a discussion of instrumental uncertainties on the Web.

Probe-Related Image Distortions. At very high magnifications and high-relief sample surfaces, the mode of imaging and the geometry of the probe tip can influence the scanned image.  Knowledge of the probe geometry then becomes important for interpretation of the image.

Suppliers of Scanning Probe Microscopes

These instruments are commercially available from several competing firms, including: 
Burleigh Instruments, Inc., Fishers, NY 
Digital Instruments, Santa Barbara, CA
TM Microscopes, formerly ThermoMicroscopes, Inc., Sunnyvale, CA
    now together form the Veeco Metrology Group.

JPK Instruments AG, Berlin, Germany

Molecular Devices and Tools for Nanotechnology, Moscow, Russia.

Molecular Imaging Corporation, Phoenix, AZ.

Omicron Vacuumphysik GmbH, Taunusstein, Germany

Quesant Instruments, Agoura Hills, CA

RHK Technology, Inc., Rochester Hills, MI

Surface Imaging Systems GmbH, Herzogenrath, Germany

Triple-O Microscopy GmbH, Potsdam, Germany

Park Scientific Instruments and TopoMetrix Corporation  have merged with to form ThermoMicroscopes.

Many of these firms offer general-purpose instruments that can handle a variety of imaging modes, while others are more specialized.  A number of physical scientists have built their own SPM instruments. The Homebrew STM Page  provides a resource for those who wish to construct their own STM.

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Footnotes, References and Links

Inventors.  The first scanning probe microscope was the scanning tunneling microscope (STM) of Binnig and Rohrer (Binnig, G., Rohrer, H., et al., (1982) Phys. Rev. Lett., 49:57.).  Gerd Binnig and Heinrich Rohrer were awarded half of the 1986 Nobel Laureate in Physics  for their design of the scanning tunneling microscope.  Ivan Amato's 1997 article, "Atomic Imaging: Candid Cameras for the Nanoworld" (Science 276(5321):1982-1985), entertainingly recounts the history of STM and AFM development.   The article is available on-line to Science subscribers.  A history of microscopy (Microscopy for Nanotechnologists by C. David  Eagle) provides perspective on the explosive development of this field.

Scanning.   The probe (or the sample under a stationary probe) generally is moved by a
piezoelectric tube.   Such scanners are designed to be moved precisely in any of the three
perpendicular axes (x,y,z).  By following a raster pattern, the sensor data forms an image of the
probe-surface interaction.  Feedback from the sensor is used to maintain the probe at a constant
force or distance from the object surface.  For atomic force microscopy the sensor is a
position-sensitive photodetector that records the angle of reflection from a laser bean focused on the
top of the cantilever.

AFM systems detect the z-displacement of the cantilever by the reflection of a laser beam focused on the top surface of the cantilever.  The feedback from this sensor maintains the probe at a constant force.

STM systems measure the quantum tunnelling current between a wire or metal-coated silicon tip and the object surface.  An electronic feedback system maintains a constant current by positioning the tip to exactly contact the surface.

NSOM systems scan an optical fiber probe over the sample. The probe has an opaque material covering its surface, except for a small aperture at the tip.  The light (usually a laser source is used) is emitted through this aperture. Image data can be gathered in transmission, reflection or fluorescence mode.  The transmission mode provides a higher signal throughput.  It can be used with specimens that are transparent and have low or moderate light absorption, particularly biological subjects.  Reflection mode is for highly scattering and opaque samples.  The resolution of optical microscopes has been limited by the wavelength of light, in practice about 400 - 500 nm. By placing a point source of light less than that distance from the sample, NSOM improves this resolution by an order of magnitude.  An NSOM is available from ThermoMicroscopes.

Modes of operation.  The manufacturers' web sites have references and application notes that are useful in understanding the advantages and disadvantages of the various modes.  The on-line guides on the Digital Instruments website is a good examples.  The magnetically-driven cantilever system (MAC ModeTM) is specific to Molecular Imaging.

Interaction force.  The z-axis (vertical) component of the force of interaction is calculated from the z-displacement of the cantilever and the spring constant of the cantilever.  From Hooke's Law,  F = - kz, where k is the spring constant.  The spring constant for a cantilever is provided by the cantilever supplier or can be determined by the investigator.  A constant force on the probe tip is maintained by feedback from measurement of the interaction force.  The probe is moved up and down to maintain the measured constant force.
Tapping modeTM, a trademark of Digital Instruments.  Tapping Mode imaging is implemented in ambient air by oscillating the cantilever assembly at or near the cantilever’s resonant frequency using a piezoelectric crystal..   To image in fluids, the entire fluid cell is oscillated to drive the cantilever into oscillation.

Tip Selection.  AFM tips are generally made of silicon or silicon nitride.  For most applications, pyramidal silicon nitride tips are used.  They are relatively durable and present a hydrophobic surface to the sample.   Conical silicon tips are often used for bio-molecular applications because they are very sharp and present a hydrophilic surface.  However, they are relatively less durable.  For the ultimate sharpness, tips of carbon nanotubes have been made.  The Rice group also has a tutorial for mounting carbon nanotube tips on commercial cantilevers.  In other cases selective modification of silicon nitride tips has been used to provide for measurement of specific molecular interactions.  STM tips are made of mechanically-formed or electrochemically-etched wire, usually noble metals or tungsten.  Digital Instruments has a useful Tip Selection Guide.   Some very high aspect-ratio tips are available from ThermoMicroscopes.  Tips are available from many suppliers.

The Effect of Instrumental Uncertainties on AFM Indentation Measurements.(1998) Mark VanLandingham,  Materials Science Program and Center for Composite Materials,
University of Delaware.

Mervyn Miles recently published a useful overview of AFM technology and applications, Scanning Probe Microscopy:  Probing the Future (1997, Science 277(5333):1845).  The article is available on-line to Science subscribers.

Sean Morgan's Scanning Probe Microscopy Page offers many links to galleries of micrographs and the home pages of SPM/STM users.

Zhifeng Shao (University of Virginia) has developed a cryo-AFM for applications in structural biology. His work on GroES, a protein chaperon, is particularly interesting.

Bibliography of Biological SPM Research (pdf file).

Course Syllabus, atomic force microscopy to undergraduates by Nancy Burnham, Worcester Polytechnic Institute, Worcester, MA.

Sample preparation is of great importance in SPM as in other areas of microscopy.  SPM analysis of biological macromolecules places particularly high demands on the quality of the substrate.  Freshly-cleaved mica surfaces has been particularly useful.  Mica presents a charged, hydrophilic surface to which proteins and other biomolecules readily bind.  Moreover, mica surfaces are nearly flat on an atomic scale and are quite clean when fresh, conditions that are ideal for scanning at high resolution.  For certain  applications, covalent attachment to the surface is be required.  A particularly useful approach has been the preparation of gold surfaces coated with protein-reactive monolayers.  Several investigators have used monolayers composed of alkanethiols and dithioalkanes. Peter Wagner (Department of Biochemistry, Stanford University) has developed a variation using N-hydroxysuccinimide ester functionalized monolayers on a gold surface.

Digital Instruments sponsors an e-mail discussion group,  "We sponsor an Internet SPM Mailing List, or "Digest," a forum for interchanging SPM-related technical information. Anyone actively involved in SPM usage is invited to join the Digest."

So.. ..who's interested in SPM?  Scientists world-wide are investigating this new technology.  Find out who's reading this page.

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Revised:  December 5, 2003
Copyright © John W. Cross