NT TR 536 Calibration Procedures For Atomic Force Microscopes (Nordtest Technical Report)

Nordtest Technical Report 536
Calibration Procedures for Atomic Force Microscopes
Anders Kühle
Danish Fundamental Metrology
Matematiktorvet 307 , DK-2800 Lyngby
Abstract:
This report documents the results of Nordtest project 1596-02 carried out by the Swedish
national metrology institute (SP), Toponova AB and the Danish Institute of Fundamental
Metrology (DFM) in year 2002. More than 8 years of research in calibration of atomic force
microscopes (AFM) has been condensed in to two procedures for calibration of metrology grade
AFM instruments. The greatest success of the project is that DFM in September 2002 has - as
one of the first laboratories in the world - successfully passed a technical assessment for using
one of these procedures for accredited (i.e. traceable) AFM measurements. It was found that the
calibration procedures in particular for the lateral plane with some limitations could also be used
for non-metrology grade AFMs if these are properly designed and linearised. In the vertical
direction this is more dubious since measurements of roughness in the 1-10 nm range gave
unacceptable differences in the results for two non-metrology grade AFMs and a metrology
AFM. The calibration procedures were successfully tested on an interference microscope and it
was shown that equivalence between AFM and interference microscope measurements can be
established through appropriate filtering, at least in the 1-10 nm range (vertical). In conclusion
there is a need to assess roughness measurement in the nanometer range with atomic force
microscopes more carefully. Also more research is required in order to establish standardised
procedures for making comparisons between AFM and interference microscope data.
Resumé:
Denne rapport gengiver resultat af Nordtest projekt nr. 1596-02 udført af Sveriges Provnings-
och Forskningsinstitut (SP), Toponova AB og Dansk Institut for Fundamental Metrologi (DFM)
i år 2002. Over 8 års forskning i kalibrering af atomic force mikroskoper (AFM) har i dette
projekt resulteret i to detaljerede procedurer for kalibrering af AFM instrumenter af metrologisk
kvalitet. Projektets største succes er, at DFM i september 2002 efter en teknisk evaluering har
opnået akkreditering til at benytte den ene af disse procedurer til sporbare AFM målinger.
Procedurerne blev også afprøvet på AFM’er af ikke metrologisk kvalitet. Resultatet er, at især
proceduren for lateral kalibrering med visse forbehold vil kunne kan anvendes på sådanne
AFM’er, såfremt disse er hensigtsmæssigt konstruerede og lineariserede. I den vertikale retning
er dette mere tvivlsomt, idet måling af ruhed i 1-10 nm intervallet gav resultater, der var
uacceptable forskelle for to ikke-metrologi AFM’er og et AFM af metrologisk kvalitet.
Procedurerne blev også afprøvet på et interferens mikroskop, og det blev bekræftet at der ved
brug af passende filtrering kan etableres korrespondance mellem ruhedsmålinger foretaget med
interferens mikroskoper og AFM instrumenter, i hvert fald i 1-10 nm området (vertikalt). Det
konkluderes, at der er behov for mere grundige undersøgelser af ruhedsmåling i nanometer
området med AFM for at kunne etablere troværdig ækvivalens mellem målinger. Der er også
behov mere forskning, der kan føre til standarder for hvordan interferens mikroskop målinger og
AFM målinger kan sammenlignes.
Danish Fundamental Metrology DFM- 02-R29 Phone 4593 1144
Matematiktorvet 307 3308 AK Fax 4593 1137
DK-2800 Kgs. Lyngby 2002 12 18 Web: www.dfm.dtu.dk
2
Contents
1 Project overview ..............................................................................................................................................3
1.1 Introduction..............................................................................................................................................3
1.2 Project background, objectives and main results .....................................................................................3
1.3 Partners ....................................................................................................................................................4
1.4 Course of the project ................................................................................................................................4
2 Lateral calibration ............................................................................................................................................5
2.1 Calibration of the metrology AFM ..........................................................................................................6
2.2 Calibration of Toponova’s standard AFM ...............................................................................................7
2.3 Calibration of SP’s standard AFM...........................................................................................................8
2.4 Calibration of Toponova’s interference microscope................................................................................9
3 Vertical calibration.........................................................................................................................................10
3.1 Calibration of the metrology AFM and of the TGZ02 grating...............................................................11
3.2 Calibration of Toponova’s and SP’s standard AFMs ............................................................................12
3.3 Measurement of the TGZ02 grating with an interference microscope...................................................14
4 Comparative roughness measurements on ball-joints for hip implants..........................................................15
4.1 AFM roughness measurements ..............................................................................................................15
4.2 Comparison between AFM and interference microscope roughness data .............................................17
5 Conclusion .....................................................................................................................................................18
6 List of equipment used...................................................................................................................................19
7 References......................................................................................................................................................19
Appendix Procedures for calibration of the three axes of an atomic force microscope using transfer standards
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1 Project overview
1.1 Introduction
Scanning Probe Microscopes (SPM) and in particular the family member called the Atomic Force Microscope
(AFM) - invented in 1986 [1] - are at present the most powerful instruments available for accurate nano- and
micrometer surface measurements including the characterisation of soft materials like polymers and biological
materials. In the AFM a sharp probe senses and follows the surface landscape while being scanned in an xy
pattern. It thereby gives a direct image of the surface area from the atomic level to about 0.1×0.1 mm2 in the
plane and up to about 6 µm for the height. The tip is an integrated part of a cantilever beam, which acts as the
force sensor in the AFM. The deflection of the cantilever, and hence the force, is most often measured by
sensing the displacement of a laser beam reflected on the back side of the cantilever on a photo detector.
Figure 1. Principle of the atomic force microscope.
Standard AFM instruments suffer from time dependent non-linearity and hysteresis in their actuating parts
which makes them difficult to use for metrology purposes. On- and off-line software linearisation schemes and
standardised instrument settings can improve this situation. The most reliable and reproducible results are,
however, achieved with metrology grade instruments, which have distance sensors on all axes. A couple of SPM
manufacturers offer a “Metrology” version of their instruments which allow for unmatched reproducibility and
linearity. Both standard AFMs and metrology AFMs require calibration in order to provide traceability to SI
units in measurements, which is the main purpose of this project.
1.2 Project background, objectives and main results

In an EU funded project “Transfer standards for calibration of scanning probe microscopes” [2], DFM has
participated in developing certified transfer standards for xy and z calibration of scanning probe microscopes.
These standards have been used in the present project to establish traceability. In the EU project, software for
evaluating SPM images of the standards and for calculating calibration parameters was developed. In the same
project and later in the EU project “Development of a basis for 3D surface roughness standards” [3] general
principles for performing the calibration have been developed. The procedures were before this Nordtest project
only documented in general terms.
Using the above mentioned transfer standards traceability to the definition of the Meter can be established in the
range of one to a few hundred micrometers in the lateral plane. In the vertical direction it is possible to calibrate
with a high accuracy down to the nanometer level. Since 1998 DFM has participated in three international
comparisons mainly between national metrology laboratories using these transfer standards.
The first goal with the project has been to formalise the calibration procedures so they become suitable for use
for accreditation purposes. Written procedures for calibration of a metrology grade AFM in all three axes has
been produced. The procedures are given in Appendix to this report. The layout the procedures follow the
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guidelines for internal quality documents at DFM and therefore have an appearance different to this report. As a
result DFM has in September 2002 successfully passed a technical assessment by the Danish accreditation body,
DANAK, which makes use of internationally acknowledged experts for such evaluations. Thus on the
background of the calibration procedures written in this Nordtest project DFM is now accredited - to our
knowledge as the first laboratory in the world - to calibrate the pitch of two dimensional gratings using AFM.
The second goal in this project was to assess how the procedures can be used for standard grade AFM
instruments and interference microscopes: what are the limitations and/or what are the requirements? The results
show that the non-linearity in the scanning devices of two standard grade AFMs used is two high for using the
procedure. However, it should be possible to linearise these instruments sufficiently well by adjusting their
internal parameters, so that the procedure is also applicable for these types of instruments – at least for lateral
calibration. The experiments showed that the procedures are highly applicable to interference microscopes.
The third goal was to compare measurements on industrial samples carried out with 1) a calibrated metrological
AFM (at DFM) and 2) two standard AFMs and 3) an interference microscope. The objective is to illustrate the
impact of the difference in the nature and quality of the instruments. Before comparing the measurements the
images were calibrated. Measurements were performed on two artificial hip joint balls from different
manufacturers. In order to facilitate direct comparison, Toponova using laser writing marked the measurement
areas on the two artefacts. It proved to be difficult to compare roughness in the 1-10 nm range for the used
AFMs, which are of different types. The main cause to discrepancies should probably be found in the non-linear
properties of the scanning devices in the instruments. However, it cannot be excluded that there are other factors
at play. It was also concluded that interference microscope images and AFM images can be compared given that
the surfaces do not have topographic variations in excess of the range covered by the phase shifting mode of the
interferometer (about 200-300 nm) and when applying appropriate filters to the AFM images.
1.3 Partners
Danish Institute of Fundamental Metrology (DFM) was the co-ordinator of the project. DFM has had as the
main task of writing and evaluating the calibration procedures (Work Package 2 and 3) and has also taken part
in the comparative measurements (WP 5). Moreover DFM has analysed all the results and compiled the final
report (WP 6). The staffs involved at DFM have been Dr. Jørgen Garnæs and Dr. Anders Kühle.
Swedish National Testing and Research Institute (SP) is the Swedish national metrology institute. SP’s role
was to evaluate the calibration procedures made by DFM (WP 3) and to perform calibration measurements with
a standard AFM instrument (WP 4). SP also measured the industrial samples provided by Toponova for the
common roughness measurement comparison (WP 5). SP’s staff has been Dr. Mikael Frennberg.
Toponova AB is a new small company which offers services and products related to surface roughness
measurement. Toponova has supplied the specimens for the comparative roughness measurements – two
artificial hip ball-joints from different manufacturers. With both a standard AFM and an interference
microscope, Toponova has measured the set of calibration standards and the hip joints. The staff involved at
Toponova has been Stefan Rosén and Dr. Bengt-Göran Rosén who is also lecturing at Halmstad Högskola.
1.4 Course of the project

Due to other business DFM started the project 3 months late according to the original schedule shown in Figure
2. Further, the work with the calibration procedures turned out to be more comprehensive than foreseen. This
also applies for the evaluation of the measured data by all partners. This resulted in a heavy workload in the
second half year of the project and a one month delay in the delivery of the final report. However, as described
above Two meetings were held, one at DFM, Lyngby, in April 2002 and one at Toponova, in May 2002.
2002
WP Work Package Name Partners Dec Jan Feb Mar Apr May Jun Jul Aug Sep Oct Nov
Phase 1 Phase 2 Phase3
1 Coordination DFM
2 Writing of detailed calibration procedures DFM
3 Test and evaluation of procedures SP, DFM
4 Test on standard AFM (non metrology grade) SP
5 Comparative measurements of test specimens TPN, SP, DFM

6 Reporting DFM, SP, TPN
Figure 2. Initial project schedule. X = Milestones.
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2 Lateral calibration
For lateral calibration a calibration grid produced within the EU project referred to in the introduction were
circulated between the partners. It was calibrated by DFM using the lateral calibration procedure described in
the appendix using a reference standard (of the same type), which had previously been calibrated and certified
by the National Physical Laboratory, NPL, in the United Kingdom. The general design of the standards is
described in the procedure (appendix), but in short they consist of a two dimensional grid etched in a silicon
substrate. The grid is described by its unit cell in terms of the side lengths, La and Lb of the unit cell and the
angle, γ, between the two corresponding component vectors. The parameters of the calibration grid are given in
Figure 3 together with a photograph of the artefact.
The calibration procedure yields three correction parameters, describing the microscope: Cx, Cy, and Cxy. The
metric coordinate system’s x-axis can be chosen so that it is parallel to the microscope coordinate system’s x*-
axis and the linear transformation can then be written as
x  C x C xy  x* 
  =   * 
 
y   0 C y  y 
The matrix elements can be interpreted as a correction parameter Cx for the x-direction, a correction parameter
Cy for the y-direction and a coupling term Cxy between the scanned x and y axes. If the x*- and y*-axes in the
microscope coordinate system are perpendicular, then Cxy = 0, Cx will be the scaling in the x-direction and Cy
will be the scaling factor in the y-direction. The angle between the x*- and the y*-direction in the microscope
coordinate system is equal to cot-1(Cxy/ Cy) for Cxy ≠ 0.
The procedure for lateral calibration has been developed for a metrology AFM, for which the non-linearity
(which is small) can be neglected in the uncertainty analysis. The procedure is designed for allowing subsequent
pitch measurements on grids with unknown pitch. For pitch measurements a small non-linearity can be
neglected when the same image size is always used both for the calibration of the AFM and for the measurement
of the grid with the unknown pitch. This is due to the fact that the calibration gives the correction factors for the
entire image – side to side, top to bottom. So as long one measures an average quantity over the entire image the
result is only affected by non-linearity if (1) the non-linearity prohibits detection of unit cells correctly (2) if the
non-linearity is so high that the inclusion/exclusions of unit cells at the boundary of the images will significantly
influence the average value.
Standard Ibsen 2D1000, S/N A00051

Quantity Value and uncertainty b
(standard uncertainty multiplied
by a coverage factor of 2) a
La (1000.3 ± 1.8) nm
Lb (999.8 ± 1.7) nm
γ (90.01 ± 0.82) deg.
Figure 3. Right: image of the calibration grid used. The chip is made from Silicon and is about 6 mm wide. The
grid itself is in the square shaped area in the middle. Left: The calibrated lengths of the unit cell vectors of the
grid, and the angle between the unit cell vectors.
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2.1 Calibration of the metrology AFM

The standard is measured with the AFM according to the procedure. The image in Figure 4 shows the result of a
60 × 60 µm2 scan. For details on how the average unit cell is extracted from the image, see the procedure
(appendix). In short, first Fourier analysis finds a rough estimate of the unit cell. Thereafter all positions of the
estimate in the image are detected. From these positions an accurate average can be calculated. Moreover the
unit cell positions can be used for evaluating the non-linearity of the instrument. The results are shown in Figure
4. The two graphs below the images show the deviation in observed unit cell positions (x, and y, respectively) in
the image from the expected positions using the calculated average unit cell. The errors are given in units of
pixels; the image has 512 pixels in both directions. The random scatter in the position errors are due to
limitations in resolution, detection
accuracy etc. accounted for in the Cx ± U(k=2) Cy ± U(k=2) Cxy ± U(k=2)
procedure (appendix). Any systematic
trend is due to non-linearity – or thermal 1.0001 ± 0.0038 1.0002 ± 0.011 0.002 ± 0.030
drift. It is seen that the peak to peak error
is less than 0.2 pixels, i.e. less than 0.4 ‰. Table 1. Results from the calibration of the metrology AFM
The calculated correction parameters and using the calibration standard in Figure 3. The uncertainties
the corresponding expanded standard are the standard uncertainties multiplied by a coverage factor
uncertainties (k = 2, 95 confidence level) of 2, for a 95% confidence level.
ae given in Table 1.
Figure 4. 60 × 60 µm2 (512×512 pixels) AFM image of the calibration grid using the metrology AFM. The two
graphs show the deviation (in x and y respectively) from the predicted positions of the unit cells in the image.
The unit cell vector components found by the procedure are: a = (999.86 nm, -28.019 nm), b = (26.027 nm,
999.17 nm).
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2.2 Calibration of Toponova’s standard AFM

The results are shown in Figure 5. It is seen this standard AFM (a DME DualScope) is highly non-linear
compared to the metrology AFM. In the y direction the non linearity exceeds the nominal unit cell size, which is
why the curve in the graph for the y-error is discontinuous around pixel position 50 (corresponding to the top of
the image). This means that the procedure for calculating the unit cell will not yield the true average, but will be
biased by the about 10% (50/512) of the detected unit cells which have been offset in the calculation. The
difference between maximum and minimum (taking into account discontinuities) of each graph gives the
maximum error in measured distance between two points in the image along the x and y axis, respectively. In
relative numbers the non-linearity is about 0.68 % in x, and 2.7 % in y. The correction parameters have been
calculated, see Table 2. They are, as mentioned above, influenced by the high non-linearity in y causing the
discontinuity in the error graph. The influence will be most significant for Cy. Because of the non-linearity and
due to lack of knowledge of the stability of the
instruments it has not been possible to estimate
uncertainties on the parameters. It is observed Cx Cy Cxy
that x-scale of the AFM image should be 1.0474 1.104 0.00243
increased with about 5% and the y-scale with
about 10%. The axes appear to be reasonably Table 2. Results from the calibration of the Toponova’s
orthogonal (Cxy ≈ 0). standard AFM using the calibration standard in Figure 3.
Figure 5. 30 × 30 µm2 (512×512 pixels) AFM image of the calibration grid using Toponova’s standard AFM (a
DME DualScope). The two graphs show the deviation (in x and y respectively) from the predicted positions of
the unit cells in the image. It is seen that the AFM is non-linear – in particular in the y direction. The unit cell
vector components found by the procedure are: a = (954.97 nm, -10.967 nm), b = (-13.825 nm, 905.63 nm).
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2.3 Calibration of SP’s standard AFM.

The comments to the calibration of Toponova’s AFM also apply to SP’s AFM. This is not so surprising since
the AFMs are from the same manufacturer. The calculated non-linearities are 0.84 % in the x-direction and 2.6%
in the y-direction. The correction parameters are given in Table 3. Precaution should be taken due to the
discontinuity in unit cell positions in the y-
direction (10% of the detected cell positions are
Cx Cy Cxy
offset by one unit cell). Again, since Cxy is small
it can be concluded that the AFM is scanning 1.0218 1.1033 0.00594
reasonably orthogonally. The x-scale of the AFM
image should be increased with about 2% and the Table 3. Results from the calibration of SP’s
y-scale with about 10%. standard AFM using the calibration standard in Figure
3
Figure 6. 40 × 40 µm2 (512×512 pixels) AFM image of the calibration grid using SP’s standard AFM (a DME
DualScope). The two graphs show the deviation (in x and y respectively) from the predicted positions of the unit
cells in the image. It is seen that the AFM is non-linear – in particular in the y direction. The unit cell vector
components found by the procedure are: a = (978.99 nm, 6.2481 nm), b = (-1.6432 nm, 906.18 nm).
The strong non-linearity in y for the two standard AFMs is caused by a combination of hysteresis and creep in
the piezo electric elements, which perform the tip motion in the microscopes. It is a commonly known
phenomenon, which can be reduced dramatically and accounted for if instead of returning to the upper left
corner after every scan down, the tip would instead reverse and scan from the bottom of the image and up.
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2.4 Calibration of Toponova’s interference microscope.

The resolution in the lateral plane of optical microscopes is ultimately limited to half the wavelength of the light
used (the Rayleigh limit). For this particular microscope the wavelength used is 547 nm. The sampling distance
is 0.22 µm in x and 0.26 µm in y. Imaging a grid with a pitch of 1 micron or less is therefore not expected to
give good contrast. However, the image in Figure 7 reveals the periodic structure of the grid reasonably well
although the modulation (top to bottom) is only about 10 nm, where the real height is about 90 nm. There is no
problem in detecting the unit cell. The graphs in Figure 7 show that the microscope, besides some edge effects,
is extremely linear. There is a clear scatter in the points, but this is due to the accuracy in the unit cell detection
algorithm when operating on an image with low sampling
density and with a poor contrast. When neglecting edge effects
and considering only the running average in terms of a fitted 5th Cx Cy Cxy
order polynomial the non linearity amounts to 0.55% in x and
0.9918 0.9963 0.000522
0.35% in y. The calculated correction parameters show that the
scales in x and y are overestimated by only 0.8% and 0.4%,
respectively, and that the axes are completely (within the Table 4. Results from the calibration of
uncertainty of the calibration of the grid) orthogonal. Toponova’s interference microscope.
Figure 7. 39 × 39 µm2 (180×152 pixels) image of the calibration grid using Toponovas’s interference
microscope (MicroXAM). The two graphs show the deviation (in x and y respectively) from the predicted
positions of the unit cells in the image. It is seen that the interferometer is highly linear. The unit cell vector
components found by the procedure are: a = (1008.1 nm, 7.1933 nm), b = (6.8768 nm, 1004.0 nm).
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3 Vertical calibration
The calibration procedure in the appendix yields three correction parameters: Czx*2, Czz*2, and Czz*. The latter,
Czz*, is the scaling (or calibration) factor of the z axis scale of the microscope at x, y, z ≈ 0 nm, Czz*2 quantifies
the weak dependence of Czz* on z to order one, and Czx*2 represents the coupling between the z-axis and the x-
axis, i.e. the image bow. The coupling between the z- and y-axes is not taken in to account since most often
images have to be levelled line-wise and thus any image bow along the y-axis will be flattened out. This implies
that step heights should always be measured with the x-axis perpendicular to the step (see Figure 8 - Figure 11).
Czx*2 is used to correct the images for the bow caused by the coupling between the x- and z-axes. Besides this,
neither Czx*2 or Czz*2 are used directly for the calculation of Czz*, but only play a role in the uncertainty budget.
The procedure for vertical calibration has been developed for a metrology AFM, for which the sensed tip
position and the physical tip position in the vertical direction is a single valued function, i.e. there is no
hysteresis. For the calibration of the metrology AFM a certified reference standard with a nominal step height
(groove) of 800 nm is used. Using the traceability provided by this standard the MDT-TGZ02 grating from
µMasch used for this project was calibrated.
Standard Nanosensors H800, SN

01.03.04
Quantity Value and uncertainty
(standard uncertainty multiplied
by a coverage factor of 2)
href (759.7 ± 8.2) nm
Figure 8. The calibrated depth of the grooves of the standard. The groove is approximately 28 µm wide. Right:
AFM image (3D projection) of the groove.
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3.1 Calibration of the metrology AFM and of the TGZ02 grating.

The Metrology AFM was calibrated using the standard shown in Figure 8. The result was:
Czx*2 = (3.9 ± 1.1) m-1 Czz*2 = (0.0035 ± 0.001) µm-1 Czz* = 1.058 ± 0.011
The uncertainties are expanded standard uncertainties (k=2). Note that the parameter Czz*2 is not to be used
directly, but is used to estimate the uncertainty when calibrating an object with unknown height: Assuming that
calibrations are always carried out such that the mean z-value is within the ±1µm range, the maximum error on
Czz* due to a height dependent sensitivity will be 1 µm × 0.0035 µm-1 = 0.0035 (brick-wall) corresponding to a
standard uncertainty of 0.0020.
An AFM image acquired near the centre of the TGZ02 grating used for circulating between partners is shown in
Figure 9. The average step height within the imaged area was derived by first calculating the average line profile
in the x-direction. Then the procedure (appendix), which is in agreement with the ISO5436 standard, was used to
measure the depth of the grooves, as indicated by the profile shown in the figure. The observed step-height is
then corrected with Czz* (above), and the uncertainty evaluated using the principles above. The result is hTGZ02 =
(104.1 ± 1.2) nm.
Figure 9. 20× 20 µm2 (512×512 pixels). Corrected image and average profile of the TGZ02 grating acquired
with the metrology AFM at DFM. The groove depth is an average over 6 parallel grooves each evaluated
according to the ISO 5436 guideline and corrected according to the procedure. The result is (104.1 ± 1.2) nm.
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3.2 Calibration of Toponova’s and SP’s standard AFMs

Images of 20× 20 µm2 were measured on the TGZ02 grating by Toponova and SP, see Figure 10 and Figure 11.
Note: The typical fingerprint of hysteresis in standard AFMs is in the profile graphs in these figures seen as
overshoots on the left side of the lines (scanning from left to right) and as dips in the left side of the grooves.
From these measurements the calibration factor Czz* was calculated, see Table 5. Because of poor knowledge of
the hysteresis, non-linearity and coupling between axes of the two standard AFMs, it is not possible to state an
uncertainty on the correction parameters. However, from previous experience with standard AFMs an estimate
of ±10% of the measurement will
cover most cases. Often it will be
much smaller – in particular if Partner Measured mean height (nm) Czz*
measurements are performed in the DFM Calibrated with certified standard 1.058 ± 0.011
same range in which the instrument is
calibrated and with the same offset in Toponova 102.4 1.017
z. In fact this can be measured as SP 114.4 0.912
described in the procedure in the
appendix. Table 5. Results from z-calibration of partner’s microscopes. Note
that the DFM’s metrology AFM is in fact not very well calibrated by
the manufacturer.
Figure 10. 20× 20 µm2 (512×512 pixels). Levelled image and average profile of the TGZ02 grating acquired
with the DualScope at Toponova. The groove depth is an average over 6 parallel grooves each evaluated
according to the ISO 5436 guideline. The result is 102.4 nm. The hysterises is seen as overshoots on the left side
of the lines (scanning from left to right) and as dips in the left side of the grooves.
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Figure 11. 20× 20 µm2 (512×512 pixels). Levelled image and average profile of the TGZ02 grating acquired
with the DualScope (standard AFM) at SP. The groove depth is an average over 6 parallel grooves each
evaluated according to the ISO 5436 guideline. The result is 114.4 nm. The hysterises is seen as overshoots on
the left side of the lines (scanning from left to right) and as dips in the left side of the grooves. Due to the slight
angle of the pattern of the grating to the xy-axes of the AFM this is not as clear as in Figure 10.
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3.3 Measurement of the TGZ02 grating with an interference microscope.

As can be seen in Figure 12 the interference microscope (see instrument list in section 6) images the rectangular
line structure as very rounded, almost as a sine wave with an amplitude less than the actual step height of
104.1 nm. This is well known to be the effect of the limited resolution, basically caused by the diffraction of
light in combination with the properties of the optics in the instrument [4]. In fact, the effective cut-off
wavelength of the instrument can be assessed by applying a Gaussian filter to an artificial image of the structure
[4] (or an AFM image). The cut-off wavelength of the filter is increased until the amplitude of the resulting
image is the same as for the interferometer image. By performing this procedure on the AFM image from Figure
9 we arrive at a cut-off wavelength of the Gaussian filter of 2.6 µm. This number will be used in section 4.2. The
resolution of the image from an interference microscope is ultimately given by the intensity in the Fraunhofer
diffraction pattern due to the limited size of the optical aperture. By comparing such optical intensity
distributions with the Gaussian weighting distribution [5] it is found that a cut-off of 2.6 µm is equivalent to an
optical resolution of approximately 1.2 µm which is somewhat more than the expected optical resolution of the
microscope (see section 6). There may be many reasons for this, but it is beyond the purpose of this report to
discuss this thoroughly.
Figure 12. 20× 20 µm2 (92×78 pixels). Levelled image and average profile of the TGZ02 grating acquired with
the MicroXAM interferometer at Toponova. Due to the limited resolution of the interferometer the lines are not
imaged as rectangular features but appear rounded. In fact they appear so rounded that the peak to valley height
is only about 80 nm – which should be compared to the expected height of 104.1 nm.
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4 Comparative roughness measurements on ball-joints for hip implants

For the comparative roughness measurements Toponova produced two measurement objects which were both
(artificial) metal hip joint balls, but from two different (anonymous) manufacturers. The two artefacts carry the
IDs “1DP” and “2BM”. The measurement areas were marked by laser writing, so that the exact same areas could
be measured by each partner.
4.1 AFM roughness measurements

AFM images acquired by the partners of the two joint balls are shown in Figure 13. The raw data has been
levelled line wise using second order polynomials in order to remove form and the influence of drift. Further the
three axes in all images have been calibrated using the calibration constants found for each microscope in
sections 2 and 3. It is seen that the two joint balls have completely different surface topography, which also
shows in the calculated roughness presented in Table 6. The two most commonly used roughness parameters Sa
(the average numerical deviation from the mean) and Sq (the standard deviation) have been calculated. It is seen
that the images from SP exhibit a characteristic waviness not present on the other images. This is due to a
commonly observed phenomenon caused by interference between the laser light reflected by the cantilever of
the AFM and by the sample surface. In order not to include this waviness in the roughness calculations all
images have been high pass filtered with a two dimensional Gaussian filter with a cut-off wavelength of
3000 nm. Otherwise a longer cut-off (e.g. 6000 nm) would have been used. Before this filtering, the images
have been low pass filtered with a Gaussian with a cut-off wavelength of 200 nm to account for the use of
different tips and for the different sampling distance in the images (512 pixels for DFM, 256 pixels for SP and
Toponova).
Partner Hip joint 1DP Hip joint 2BM

Sa (nm) Sq (nm) Sa (nm) Sq (nm)
DFM 2.78 8.69 1.10 1.54
Toponova (AFM) 5.61 11.6 2.10 2.88
SP 3.43 6.52 1.51 2.07
Table 6. Summary of the roughness data calculated for the images in Figure 13.
It is seen that though the roughness values are within the same range they do not coincide very will with up to
almost a factor of two between the smallest values and the highest values. It is difficult to point at one
straightforward explanation, especially because the measurements of the vertical calibration grating gave results,
which agreed within about 10%. We have identified three possible causes to the discrepancies:
1) Deviations within 1-2 nanometers can be explained by the fact that all AFMs used in this project are
operating in dynamic mode, i.e. using the cantilever as a resonator. It is well known that this mode of
operation is sensitive to adsorbed water layers and other contamination on the surfaces, which the tip may
track instead of the real surface. But as the images do all have a comparable resolution in xy (showing no
blur due to contamination) this is probably not the cause.
2) In the dynamic mode the cantilever sometimes is un-stable switching between attractive and repulsive
modes [6]. When this is happening fast it is difficult to see this as other than random noise in the images –
increasing the roughness with up to 1-5 nm. Another cause to increased roughness can simply be
mechanical or electrical noise in the microscopes.
3) When comparing profiles taken over specific holes on the sample “1DP” for the three microscopes we
observe that the measured depth of the holes actually do vary from microscope to microscope in the same
way as the roughness data in Table 6. This first of all excludes the possibility that it is the image processing
(filtering) which disturbs the data. It also shows that the calibration performed with the 104.1 nm grating is
not very useful. This is most probable due to the non-linearity and hysteresis of the piezoelectric scanners in
the two standard AFMs.
In conclusion we have found if difficult to compare roughness in the 1-10 nm range for AFMs of different types.
The main cause to discrepancies should probably be found in the non-linear properties of the scanning devices
in the instruments of the standard type. However, it cannot be excluded that there are other factors at play.
3308 AK 2002 10 29
16
Hip joint ball 1DP Hip joint ball 2BM
DFM
Topo-
nova
SP
Figure 13. AFM images, of the two artificial hip joint balls, named “1DP” (left), and “2BM” (right), taken with
the AFMs at the three partners; top: DFM, middle: Toponova, bottom: SP.
3308 AK 2002 10 29
17
4.2 Comparison between AFM and interference microscope roughness data

Interference microscope images of the two artefacts were made by Toponova. The images were 2-3 times larger
than the corresponding AFM images. In order to make comparison easy, the regions also imaged by AFM have
been identified and have been cut out of the original interference microscope images. The results are shown in
the right column of Figure 14. It should be easy to make the association to the AFM images in Figure 13. In
order to compare roughness and visual resemblance the AFM images generated by the metrology AFM (top of
Figure 13) have been low pass filtered with a two dimensional Gaussian filter with a cut-off wavelength of
2.6 µm as described in section 3.3. The resulting images are shown in the left column of Figure 14. The visual
resemblance is striking, as also observed in recently published work [5]. The roughness calculated from the
images with out further filtering is shown in Table 7. The data from the “2BM” artefact coincide almost
completely. The roughness values for the “1DP” artefact are equal within about 30% - a rather large difference.
The highest values are for the AFM. The difference may be explained by the fact that the surface of the “1DP”
artefact has topographic variations (deep holes) in excess of what the interference microscope can handle when
operating in phase shifting mode, which is the mode used here. In fact when zooming in on the holes we see that
many of them have strange (non-physical) structures at the bottom and are less than 1/3 as deep as in the AFM
images.
Filtered AFM Interference microscope
Figure 14. Filtered AFM images (left) and interference microscope images (right) of the two hip joint balls.
Top: 1DP, bottom: 2BM. The AFM images were low pass filtered with a two-dimensional Gaussian envelope
with a cut-off wavelength of 2600 nm.
3308 AK 2002 10 29
18
In conclusion (supported by [5]) we can say that interference microscope images and AFM images can be
compared given that the surfaces do not have topographic variations in excess of the range covered by the phase
shifting mode of the interferometer (about 200-300 nm). We also conclude that the method proposed in [4] and
used in section 3.3 for determining the effective cut-off wavelength of the interference microscope within the
framework of Gaussian weighting functions is useful.
Partner Hip joint 1DP Hip joint 2BM

Sa (nm) Sq (nm) Sa (nm) Sq (nm)
DFM 7.98 17.5 0.96 1.22
Toponova (interferometer) 6.50 12.4 0.96 1.21
Table 7. Summary of the roughness data calculated for the AFM and interference microscope
images in Figure 14.
5 Conclusion
In this project more than 8 years of research in calibration of atomic force microscopes has been summarised
and brought in to the form of two procedures for calibration of metrology grade AFM instruments: one for the
lateral plane and one for the vertical axis. The procedures include a full uncertainty analysis. As a result DFM
has in September 2002 successfully passed a technical assessment by the Danish accreditation body, DANAK,
for using the lateral calibration procedure to calibrating the pitch of two dimensional transfer standards for
clients. To our knowledge this is one of, if not the first accredited use of AFM in the world. The challenge for
the future is to disseminate traceable measurements in the nanometer scale to industry and to extend the
catalogue of traceable measurement types through research in targeted areas as f. ex. micro optics.
It was investigated how the calibration procedures work for standard AFMs and interference microscopes. The
conclusion is that the non-linearity in the scanning devices of the two standard grade AFMs used is two high for
using the procedure for lateral calibration. A better software linearisation and better control of the piezo electric
scanners in the instruments could change this. The vertical calibration procedure could be applied, but
unfortunately it was not found useful as explained below. The experiments showed that the procedures are
highly applicable to interference microscopes, only the grating used for vertical calibration should have a higher
pitch than the one used, preferably about 20 µm.
Roughness measurements (1-10 nm range) on two artificial hip joint balls with three AFMs did not give data
which were satisfactory consistent, even though the vertical axes of the instruments were calibrated in the
100 nm range. The cause should probably be found in the non-linear and hysteretic nature of the piezo electric
scanners in the two standard AFMs. But no definite cause was identified. In order to understand this better and
establish equivalence there is thus a need to assess roughness measurement in the nanometer range with atomic
force microscopes more carefully.
Last it was confirmed that metrology AFM and interference microscope roughness measurements can be
compared when applying Gaussian filters to the AFM images given that the surfaces have small topographic
variations, e.g. less than 50 nm. The results indicate that the properties of the filter can be determined from
images of rectangular structures. These conclusions are very promising, however, more work is required in this
field in order to establish standardised procedures for making comparisons.
3308 AK 2002 10 29
19
6 List of equipment used

Metrology AFM (DFM) Digital Instruments DI3100 with metrology head, 70µm×70µm×6µm
measurement volume.
Standard AFM (Toponova) Danish Micro Engineering, DualScope 45-40 AFM, 40µm×40µm×2.7µm
measurement volume.
Standard AFM (SP) Danish Micro Engineering, DualScope 45-40 AFM, 40µm×40µm×2.7µm
measurement volume..
Interference microscope ADE Phase Shift, MicroXAM 100 HR, 50x objective, CCD sampling
(Toponova) 0.22 µm×0.26 µm, optical sampling 0.5 µm×0.5 µm, wavelength 547 nm.
7 References
[1] Atomic Force Microscope, G. Binnig, C.F. Quate, CH. Gerber, Phys. Rev. Lett. 56, 930 (1986).
[2] SMT4/CT95/2018.
[3] “SURFSTAND”, SMT4/CT98/2209.
[4] W. Hillmann, U. Brand, M. Krystek, Measurement 19 (2), 95-102 (1996).
[5] J. Garnaes, N. Kofod, A. Kühle, C. Nielsen., K. Dirscherl, L. Blunt, Precision Engineering 27 (1), 91-98
(2003).
[6] A. Kühle, A. H. Sørensen, J. B. Zandbergen, and Jakob Bohr, Appl. Phys. A 66, 329-332 (1998).
3308 AK 2002 10 29
Page 1 of 31
2002-12-18
Appendix to
Nordtest Technical Report 536

Calibration Procedures for Atomic Force Microscopes
Procedures for the calibration of the three axes of an
atomic force microscope by the use of transfer standards

AFM calibration procedures Page 2 of 31
Contents
1. Objective ..................................................... 3
2. Description of equipment ...................................... 4
2.1 The microscope ................................................ 4
2.2 Clean room bench .............................................. 4
2.3 Software ...................................................... 5
2.4 The two-dimensional standard .................................. 5
2.5 The step height standard ...................................... 6
3. Lateral calibration using transfer standards .................. 7
3.1 Theory for lateral calibration ................................ 7
3.2 Measurement of reference standard ............................ 12
3.3 Image processing ............................................. 13
3.4 The certificate .............................................. 14
3.5 Uncertainty analysis ......................................... 14
3.6 Uncertainty budget ........................................... 18
4. Vertical calibration using transfer standards ................ 19
4.1 Theory for vertical calibration .............................. 19
4.2 Measurement of reference standard ............................ 24
4.3 Image processing and calculations ............................ 25
4.4 The certificate .............................................. 26
4.5 Uncertainty analysis ......................................... 26
5. Addendum - Estimation of a reference plane, levelling ........ 31
1. Objective
The objective of this calibration procedure is to calibrate a metrology
atomic force microscope (AFM), in units of metre,
1. in the lateral plane for a nominal scan area of 60 µm x 60 µm.
2. in the vertical direction for a nominal length up to 6 µm
at a reference temperature of Tref = (23º± 2)°C and a relative humidity of
Href = (50 ± 10)%,
The lateral calibration is based on a two dimensional grid with a cali-
brated pitch in the x and y direction. The vertical calibration is based on
a calibrated step height and a flat surface. The result of the vertical
calibration is a 2 x 2 correction matrix and for the vertical calibration
it is a set of three correction parameters. Using these corrections a re-
corded image can be described in a metric coordinate system off-line.
The equipment is described in section 2, the theory for the lateral cali-
bration is presented in section 3 and the theory for the vertical calibra-
tion in section 4.
1.1.1 The metric coordinate system

Let (z, x, y) be an orthogonal coordinate system with the unit of one metre
on the axes. This coordinate system will be referred to as the “metric co-
ordinate system” and no superscript will indicate that the basis for a co-
ordinate set is the metric coordinate system.
The certified parameters of the transfer standards are given in the metric
coordinate system.
2. Description of equipment
2.1 The microscope

A commercial AFM with a metrology head (scanner) [7] is used (see Figure
1). The z-height is sampled at a constant time interval as a function of
the approximately orthogonal x and y directions. The constant lateral scan-
ning speed is obtained and controlled by a closed loop based on feedback
from capacitive distance sensors. The instrument has a nominal horizontal
scanning range of 70 µm x 70 µm and a nominal vertical range of 6 µm with a
maximum sampling of 512 lines of 512 pixels.
The software which control and record the image cannot assessed directly,
and the microscope including the software is calibrated as a hole.
Vertical and
lateral capac-
Parallelogram ity position
sensors
Reference cu-
be
Cantilever
and tip Sample
Figure 1 Sketch of the metrology system. The metrology frame is indicated for one
axis only. The figure is modified from [1].
During calibration it is indirectly the sensitivity of the capacitive sen-
sors which is determined. The relationship between distance and measured
capacitance may deviate from theory due to a number of causes [2]. These
are bow of capacitor plates, plate tilt (that is the plates are not 100%
parallel), and stray capacitance. Stray capacitance can be reduced by the
use of guard rings; tilt errors can be reduced by using large gaps (plate
spacing). The draw-back is a reduction in the signal to noise ratio of the
sensor. Calibrating the sensors and applying corrections to the signals can
account for most of these errors causing deviations from linearity. How-
ever, for practical capacitive sensors a small amount of non linearity is
inevitably present. For the microscope used the integral non linearity is
specified by the manufacturer to less than 1 ‰ of the scan range in the
lateral plane and 1 % in the vertical direction. For the lateral plane we
observe a non linearity less than 0.4 ‰.
2.2 Clean room bench

The microscope is place on a vibration isolation table in a clean room
bench which have a downflow of filtered air, with controlled temperature
and humidity. The bench enclose the microscope by windows.
2.3 Software
The software used (Scanning Probe Image Processor, SPIP [10]) for the de-
scribed analysis is an image processing program with special tools for ac-
curate characterisation of image structures and calibration of in particu-
lar scanning probe microscopes. A number that is stored in each data file
generated identifies the current version of the software. Every new release
of the software taken in use is validated according to ISO 17025 require-
ments and [3].For precautions and details about access to the programme a
Software Validation Checklist should be maintained.
For calculating some of the correction parameters and all the uncertainty
budget the program DFM-GUM is used. Every release of DFM-GUM is validated
according to ISO 17025 and [3] by the manufacturer. For precautions and de-
tails about access to the programme a Software Validation Checklist is
maintained.
2.4 The two-dimensional standard

The transfer standard to be used consists of a two dimensional lattice of
features (either bumps or pits) on a silicon wafer. The lattice is made by
pattering a mask-layer by holography, that is, using the beam coherence of
a laser to form an interference pattern on the photo resist. The exposed
(or non exposed) part of the resist is then removed and the pattern is sub-
sequently etched into the wafer surface. The average length La and Lb and
the angle γ between closest features defines the lattice (see Figure 2).
Nominally the two-dimensional lattice is square, however, in practice the
lattice is not perfectly quadratic but rather a little bit oblique, that is
L a ≠ L b and γ ≠ 9 0 o .
2.4.1 Traceability
The three quantities, La , Lb, and the angle γ , have been calibrated by a
diffraction technique using a helium-neon laser, calibrated against an io-
dine-stabilized reference laser at NPL [4]. The expanded uncertainties of
the three quantities are given in the calibration certificate. The artefact
has La = Lb = 895 nm and γ ≈ 91°.
Figure 2. Drawing of the

reference standard. The
lengths La and Lb are the
nearest neighbour dis-
tances between the charac-
teristic features forming
the lattice. The angle be-
tween the rows is denoted,
γ.
2.5 The step height standard

To calibrate and subdivide the z-scale a calibrated step height standard is
used. It has a nominal step height of 800 nm, a nominal width of the step
of 20 µm, and is called H800 see [5]. It is made in silicon/silicon oxide.
The surface is made homogeneous, opaque and conductive by a thin metallic
Figure 3 To the left is a part of the step height standard showing the reference
field and the positions E1 to E4 chosen for calibration. To the right a photogra-
phy of the standard with the same orientation.
film metallic film (PtIr-Cr) approximately 70 nm in thickness, see Figure

4.
For calibration the area E1 is used.
2.5.1 Traceability
The depth of the groove in the reference area was measured at four spots
named E1 to E4 (see Figure 4). The groove depth was determined by fringe
evaluation in a calibrated interference microscopy [6]. As this method al-
lows only fringe fractions to be determined, a calibrated contact stylus
instrument estimated the number of whole interference fringe.
3. Lateral calibration using transfer standards

To calibrate the microscope an image of the reference grating is recorded,
levelled, and the exact peak positions for each feature is estimated based
on maximal correlation with a single feature on the sample. Then a linear
transformation of the average peak positions into the certified lattice
constants is estimated. This transformation is defined by a correction pa-
rameter cx for the x-direction, a correction parameter cy for the y-
direction and a coupling term cxy between the scanned x- and y-axes.
3.1 Theory for lateral calibration

This chapter gives the theoretical principle for the levelling, the estima-
tion of feature positions and the estimation of the linear correction pa-
rameters and the algorithms for the software used.
3.1.1 The microscopes coordinate system

During the measurement the microscope samples a set of equidistant height
values of the surface along two approximately orthogonal directions.
The sampled coordinates (that is, the recorded image) are functions of the
environmental conditions of which the most significant are temperature T,
humidity H, and the height of the probe ∆hp. These parameters influence the
mechanical connection between the probe and the sample, the sensitivity of
the capacitive sensors and the Abbé offset. Due to relaxation, the observed
coordinates further depend on the history.
The non-linearity in the recorded image is less than 1 ‰ (in the xy plane)
over the full scan area [7]. The recorded images can therefore be treated
as linear in the analysis below.
Let i = 1, … , n, and j = 1, … , m be indices, and n, m be the number of
indices. Let the microscope be in (“perfect”) equilibrium at the reference
temperature Tref = 23ºC and a relative humidity of Href = 50%, and a refer-
ence height of the probe ∆hp,ref = 0, call the “reference condition”. At
these reference conditions the sampling points during scanning can be de-
scribed as an oblique coordinate system,
z*(c* xi, c* y j) = z*(x* i , y* j) (1)
where z*, x*i and y*j are the vertical and lateral positions in the z*, x*
and y* direction which would be displayed by the microscope at the refer-
ence conditions, with the dimension length, and where c*x and c*y are the
microscopes scaling factors, with the dimension length, used by the micro-
scope. This reference coordinate system will be referred to as the “micro-
scope coordinate system” and a “*” will indicate that the basis for a coor-
dinate set is the coordinate axes of the microscopes coordinate system. The
microscope often uses the same scaling factor, and the same number of pixel
and along the x*i and y*j directions corresponding to a square image. Cor-
rect microscope scaling factors, and a correct angle between the x* and y*
direction can be calculated from the estimated correction parameters cx cy,
and cxy , however, they are not necessary to estimated.
To simplify the notation the discrete values of the lateral position x*i
and y*j, are substituted with only x*, and y* and redefined as a continuous
variable when, for example, analysis is done at sub pixel level.
3.1.2 The actual traced coordinate system

Due to (unknown) variations in temperature, humidity and Abbé offset, the
sampling is not done (exactly) in the microscope coordinate system. The ac-
tual traced (or “true”) position during sampling is written as
z*’(x*’i, y*’j) . This oblique coordinate system is referred to as the “ac-
tual traced coordinate system” and the positions as the actual traced posi-
tions. A “ *’ “ will indicate that the basis for a coordinate set is the
coordinate axes of the actual traced coordinate system which are (slightly)
different from the microscopes axes.
In a measurement the z*’ positions are sampled at the actual traced lateral
coordinate positions (x*’i,y*’j) but they are displayed as if they were sam-
pled in the microscope coordinate system (x*, y*) in the positions
(x*i, y*j) = (c*xi, c*yj).
To simplify the notation the discrete values of the lateral position x*’i
and y*’j, are substituted with only x*’, y*’ and redefined as a continuous
variable when, for example, analysis at sub pixel level has been done.
3.1.3 The correction parameters cxx*, cyy*, and cxy*

We want to find the linear transformation matrix C of the (displayed) mi-
croscope coordinate system (x*, y*) into a metric coordinate system (x, y),
see definition in section 1.1.1.
The metric coordinate systems x-axis can be chosen so that it is parallel
to the microscope coordinate systems x*-axis and the linear transformation
can then be written as (see also [8] and [9])
x  cxx * cxy *  x* 
  =   *  ( 2 )
 
y   0 cyy *  y 
where cxx*, cyy*, and cxy* are the correction parameters to be estimated by
the calibration procedure. By choosing the x and x* direction to be paral-
lel the matrix element cyx* become zero.
The matrix elements can be interpreted as a correction parameter cxx* for
the x-direction, a correction parameter cyy* for the y-direction and a cou-
pling term cxy* between the scanned x and y axes.
If the x*- and y*-axes in the microscope coordinate system are perpendicu-
lar, then cxy* = 0, cxx* will be a scaling in the x-direction and cyy* will be
a scaling factor in the y-direction. The angle between the x*- and the y*-
direction in the microscope coordinate system is equal to cot-1(cxy*/ cyy*)
for cxy* ≠ 0.
The correction parameters are functions of same parameters as the sampled
coordinates (see section 3.1.1) The measurand during calibration is defined
as the values at equilibrium at the reference temperature Tref = 23ºC and a
relative humidity of Href = 50%, and a reference height of the probe
∆hp,ref = 0, that is
cxx * ≡ cxx *(Tref, H ref, ∆hp,ref)
( 3 )
cyy * ≡ cyy *(Tref, H ref, ∆hp,ref)
cxy * ≡ cxy *(Tref, H ref, ∆hp,ref)
For further details about influence parameters see section 3.5.

3.1.4 Plane correction

The surface plane and the x*,y* plane in the microscope coordinate system
will inevitably be tilted slightly with respect to each other. Further
drift along the z-axis may disturb the image and sometimes the tip makes
sudden jumps causing scan lines to be shifted in height. A levelling proce-
dure is therefore required, before the position of the features (see sec-
tion 3.1.5) and the step height can be estimated.
If there is no height shifts between individual scan lines a 2. order poly-
nomial plane fit is sufficient for plane correction. Otherwise line wise
levelling is performed iteratively 3-4 times using 2. order polynomia, re-
stricting the estimation volume to the basal plane1 of the image using “In-
side Color Range” option in SPIP. This in addition removes image bow. For
an exact mathematical description of the algorithms estimating the refer-
ence lines see Addendum (section 6).
The software maintains the original x* and y* scale after levelling, that
is, it displays the lateral distance projected to the corrected plane and
not the distance which would be measured on a rotated surface. To correct
for this the microscope coordinate set (x*, y*) should be transform as fol-
low
 x*   x* / cos β x 
  →  
 (4)
 y*  y* / cos β y 
 x*'  x*' / cos βx 

  →  *  (5)
 y *'  y ' / cos β y 
   
where βx and βy are the tilt angles of the surface of the standard with re-
spect to the scanning plane along the x-axis and the y-axis, respectively.
In practice the tilt angles, βx and βy, are often below 0.25°, which implies
the cosines to deviate from unity with less than 1×10-5. Therefore the
cos(βx) and cos(βy)terms are considered unity in the analysis and the asso-
ciated uncertainty vanishing. The tilt angle can be measured from the re-
corded images. For the y-axis it is estimated as the average of the up and
down scan, for the x-axis it is estimated as the average of a trace and re-
trace scan. If it is higher than 0.25° the images are corrected by multi-
plying the x- and y-dimensions with 1/cos(βx) and 1/cos(βy), respectively,
before the plane correction is applied
3.1.5 Determination of observed unit cell features

From the surface coordinates z(x*, y*) of a two dimensional grating, at
equilibrium and under reference conditions, we want to estimate the average
position of the repeated feature by estimating the position of all the fea-
tures (see Figure 4 (a)). The algorithm (called “Fine Linearity Correc-
tion”) works as follows (for further details see [10] and [11]):
1. An approximate estimate of the unit cell is done by calculating the Fou-
rier transform of the levelled image. Based on this a single feature
representative of the lattice zAu(c*x o, c*y p) with area Au and pixel co-
ordinates o, p ∈ Au is selected (see Figure 4 (b)).
1
The basal plane is defined as the bottom part of the surface in case of a continuous surface
with bumps and as the top part of the surface in the case of a continuous surface with pits.
2. The cross-correlation function CAu(x*,y*) between the particular unit

cell area Au and the surface coordinate set z(x*,y*) is calculated by
(see Figure 4 (c))
C Au(x*, y*) = ∑ z(x* + c* xo, y* + c* y p) ⋅ zAu(c* xo, c* y p) (6)

o,p
where the zero point for o and p has been chosen appropriately.
3. The peaks, p*kl, which are the local maximum in the cross correlation
function, represent positions of the unit cell in the microscope coordi-
nate system. Mathematically it is expressed as
{p* kl} = {(x*, y*) C Au(x*, y*)(x*,y*) is a local maximum} (7)

kl
where, k = 1, … K and l = 1, … L are indices for the cross-correlation

maxims (with an appropriated numbering), and where K·L is the approximate
number of features identified. Let the position error vector ekl be the
difference between the positions of peaks in the cross-correlation func-
tion and the nodes in a lattice formed by the unit vectors a*, b*, that
is,
e kl = p* kl − (p* 00+ ka*+ lb*) (8)
where p*00 is the coordinates of the highest peak in the cross-

correlation function. The average unit cell vectors a*, b* are then fit-
ted by minimising the position error, that is,
min (∑ e kl2)
a*,b*,p *00
kl (9)
In wording the co-ordinates of the average unit cell vectors a*, b* are
calculated by averaging the nearest neighbour vectors between peaks in
the cross-correlation function, see Figure 4. All positions and vectors
are identified at sub-pixel level.
0.94µm × 0.86µm
20µm × 20µm 20µm × 20µm
Recorded image Unit cell template Cross-correlation function
Figure 4. Example of an image recorded of a standard, the identified unit cell tem-
plate, and the cross-correlation function. In order for the structures to be dis-
cernable, this example is from a 20µm × 20 µm scan and not for a full range scan.
Figure 5 Result of the calculation reported by the software. The numbers named
“a vector – x”, “a vector – y”, “b vector – x” and “b vector – y” correspond to the
“observed” unit cell vector coordinates ax*' , a*' *'
y , bx
*'
and b y for the a*' and b *'
vectors used in the calculations of the correction parameters. The program can also
correct the image for non-linearity. This option is not used in this procedure. The
check box “Include Border Region” should remain unchecked. Note that the program
can not distinguish between a*' and b *' . Further the program uses a y-axis which is
-90° rotated from the x-axis. It is therefore up to the user to sort out which vec-
tor is which, and to transform the orientations into a “right handed” coordinate
system.
3.1.6 Estimation of correction parameters cxx*, cyy* and cxy*

From ( 2 ) it is seen that the average unit cell in the metric coordinate
system can be written as
 ax  c cxy *  a*x 
  =  xx *   (10)
 ay   0 cyy *  a*y 
    
 bx  c cxy *  b*x 
  =  xx *   (11)
 by   0 cyy *  b*y 
    
where cxx*, cyy* and cxy* are the unknown correction parameters to be esti-
mated.
Though it has no implication for the actual calculations, we choose by con-
vention the a* =(a*x, a*y) vector to be the one with the largest x*-
coordinate, i.e. the vector most parallel to the x*-axis.
In the metric coordinate system (x, y) the relationship between the average
length La and Lb and the angle γ between closest features on the reference
standard can be written as:
a = La b = Lb a ⋅ b = La ⋅ L b ⋅ cos(γ ). (12)
By inserting (10) and (11) into (12), a restraint between the unknown ma-
trix elements cxx*, cyy* and cxy*, and the certified dimensions La, Lb and γ is
obtained resulting in three equations with three unknowns [8, 9,12]:
A
c xx * = − (13)
*
a y ⋅ b *
x − a* x ⋅ b * y
La ⋅ Lb ⋅ sin(γ )
cyy * =
A (14)
b*x ⋅ b*y ⋅ L2a + a*x ⋅ a*y ⋅ L2b − (a*y ⋅ b*x + ax* ⋅ b*y ) ⋅ La ⋅ Lb ⋅ cos(γ )
cxy * =
A ⋅ (a*y ⋅ b *x − a*x ⋅ b*y ) (15)
where
2 2
A = b*y ⋅ L2a + a*y ⋅ L2b − 2 ⋅ a*y ⋅ b*y ⋅ La ⋅ Lb ⋅ cos(γ ) .
(16)
3.2 Measurement of reference standard

It is along the fast scanning direction (the x-direction) the best calibra-
tion of the AFM is obtained. The default scan angle is 0°, which is ap-
proximately parallel to the long dimension of the cantilever. A high accu-
racy perpendicular to the cantilever can be obtained by performing the
calibration with a scan rotation of 90°. An example of an image is shown in
Figure 4.
3.2.1 Measurement procedure

Before the measurement start the microscope should have been switched on
for at least 12 hours. The date is recorded.
The grating is placed into the microscope in the oriented as shown in
Figure 6.
Figure 6. The reference stan-

dard is aligned with the black
line (on the mounting plate)
approximately parallel to what
is the x axis of the microscope
when scanning at 0° scan rota-
tion. The lattice vector in
γ this direction is named a; the
b perpendicular lattice vector is
a x named b .
y x y
x
° °
At 90 scan rotation At 0 scan rotation
The microscope is operated in Tapping Mode according to the guidelines and

precautions described in the manual for the metrology microscope system
[13]. All experimental settings are automatically recorded and stored in
the image file.
During the recording of the image it is controlled that the trace and re-
trace are identical despite from a reasonable drift in the z direction, and
that the up and down trace are identical despite from a reasonable drift. A
reasonable drift in the lateral plane is when identical features are
shifted not more than approximately two pixel. This check also imply that
the gain settings, and force exerted on the sample by the tip does not af-
fect the image quality.
The AFM is calibrated by acquiring an image of the standard according to
Table 1. Before acquiring the calibration image, C1, the microscope should
have been scanning continuously for 1 hour or more, that is, an image of
the trace up is shown on the screen. The filename is recorded.
It is the “Height” information in the “trace” direction, which should be
saved and used for the calibration. The minimum and maximum temperature and
humidity under the calibration is recorded. In order to calibrate the in-
strument in its full measurement range the largest possible quadratic scan
area is chosen. This will usually be around 60µm × 60µm.
Table 1. Parameters for images used for calibration.
Standard Scan size Sampling Scan rota- Scan Scan

tion rate direction
Ibsen La, La 60µm × 60µm 512 × 512 0° or 90° 0.14Hz Trace down
~ 895 nm
3.3 Image processing

The algorithm in SPIP named “Fine Linearity Correction” is run (see Figure
5) and the observed average unit cell coordinates a*’x, a*’y, b*’x, and b*’y
are recorded. When SPIP has completed the algorithm SPIP saves the results
in a file named “<imagefilename>.cal”.
The model function to calculate cxx*, cyy*, and cxy* (see section 3.5.1) is
typed into DFM-GUM templates. To rule that the typing was done correct, it
is controlled that the appropriate values in the uncertainty budget in sec-

tion 3.6, gives the same results. Then the observed average unit cell coor-
dinates is typed into DFM-GUM worksheets and the calculated correction pa-
rameters and uncertainties are recorded.
3.4 The certificate

Before the certificate is written the calibration should be accepted: The
parameters cxx* and cyy* should be ≈ 1, and cxy* ≈ 0. Additionally the differ-
ences to previous values should be reasonable.
3.5 Uncertainty analysis

The input quantities to the model function are significantly influenced by:
• The accuracy of the calibration of the reference standard.
• Lack of accuracy when recording and processing the image.
• The lack of equilibrium and knowledge about the environmental condi-
tions.
• The lack of perfect movement of the probe in the microscope coordi-
nate system.
Other influence sources have been considered and found not to be signifi-
cant. These are, the horizontal orientation of the grating, the tip shape,
a small amount of dust on the sample, electromagnetic interference from the
power line, acoustical noise and the force exerted on the sample by the
tip.
The evaluation of the standard uncertainty on the input quantities is based
on the experience and experimental documentation in the laboratory summa-
rized in the scientific publications [8], and [9], the Ph.D. thesis [12]
and the report on the international supplementary key comparison NANO4
[14].
3.5.1 The model function

The most important errors during recording (to first order) is
δxM , δyM - the change in tip position due to thermal contraction
and expansion in the microscope during the imaging
relative to the side lengths of the image
δxC , δyC - the difference between the actual sensitivity of the
capacitive sensors during the measurement and the sen-
sitivity at the reference conditions
δxA , δxA - the change in sensitivity due to the Abbé offset error.
Taking into account the above mentioned error sources, the actual traced
lateral coordinates of the tip (x*’i, y*’j) can then be expressed as
 x*'  1 + δ xC 0 1 + δ xA 0 1 + δ xM /m δ xM  x* 

      * 
 y *' =  0 1 + δ yC  0 
1 + δ yA  0 1 + δ yM  y  (17)
    
 x*' 1 + δxC + δxA + δxM /m δxM  x* 

  =   * 
 y *'  0 1 + δyC + δyA + δyM  y  (18)
    
where the matrix element δxM is due to the fact that the drift will cause
the start of each line in the x-direction to shift (for further details see
[11]).
The microscope coordinate set (x*i, y*j) as function of the actual traced
coordinates can then be expressed as
 x*  1 − δxC − δxA − δxM /m − δxM  x*' 

  ≅    (19)
y 
*  0 1 − δyC − δyA − δyM  y*'
  
This imply that
 ax  C C xy 1 − δ xC − δ xA − δ xM /m − δ xM  a*'x 
  =  x   *  (20)
 ay  0 
C y  0 1 − δ yC − δ yA − δ yM  a ' 
    y 
 bx  C C xy 1 − δxC − δxA − δxM /m − δxM  b*'x 

  =  x    (21)
 by   0 C y  0 1 − δyC − δyA − δyM  b*'y 
     
and after substitution into equation (10) and (11) the original solution in
(13), (14) and (15) are now
A
cxx * ≅ −
( * * * *
)
a 'y ⋅ b 'x − a 'x ⋅ b 'y ⋅ (1 − δxM /m − δxC − δxA ) (22)
La ⋅ Lb ⋅ sin(γ)
cyy * ≅
(
A ⋅ 1 − δyM − δyC − δyA ) (23)
b*'x ⋅ b*'y ⋅ L2a + a*'x ⋅ a*'y ⋅ L2b − B ⋅ La ⋅ Lb ⋅ cos(γ)

C xy ≅ + δxM
( ) (
A ⋅ a*'y ⋅ b*'x − a*'x ⋅ b*'y ⋅ 1 − δyM − δyC − δyA ⋅ )
(24)
where
(25)
A = b *'y2 ⋅ L2a + a*'y2 ⋅ L2b − 2 ⋅ a*'y ⋅ b*'y ⋅ La ⋅ Lb ⋅ cos(γ )
B = (a*'y b*'x +a*'x b*'y ) (26)
and where
La, Lb, γ dimensions of the transfer standard, from the cer-
tificate
a*’x, a*’y, b*’x, b*’y observed average lattice vector coordinates ex-
pressed in the microscope coordinate system.
Detailed descriptions are given in separate paragraphs below.
3.5.2 The reference standard: The certified lengths and

angle (La, Lb, and γ)
The estimate is the dimensions and angle of the grating-standard given in
the certificate (La ≈ La ≈ 895 nm, γ ≈ 91.1°). The standard is manufactured
from silicon for which the thermal expansion coefficient at room tempera-
ture is about 5×10-6 K-1 and this is insignificant compared to the other lar-
ger contributions to the resulting uncertainty. Temporal drift of silicon
is in the order of 10-8 per year, which is also insignificant. Thus the un-
certainty is estimated to be the uncertainty on the calibration stated in
the certificate:
u(La) ≈ u(Lb) ≈ 0.5 nm γ ≈ 0.05°
3.5.3 Image quality and processing: The observed average

unit cell coordinates ax*' , a*' *'
y , bx and b y*'
The estimates are calculated by the image processing program [10] as func-
tion of the cross-correlation maxima in the recorded and levelled image,
see Section 3.1.5. (eqs. (6), (7), (8), and (9)).
The estimated average “recorded” unit cell coordinates are functions of and
influenced by:
A. The plane correction method ((42), (43) and (44)) and remaining tilt an-
gles βx and βy (4),
B. The number of discrete sampling points n, m (1).
C. The number of features (unit cells) detected in the image (9),
D. The algorithm that estimates the local maximum in the cross-correlation
function in (7).
E. The position errors ekl of the cross correlation peak positions, p*kl in
(8).
A) Tilt (cosine error) is accounted for as described in Section 3.1.4. The
remaining uncertainty from plane correction is denoted, up (p for Plane
correction).
B)-E) These contributions are accounted for collectively the uncertainty is
denoted ucc (cc stands for Cross Correlation unit cell detection).
The resulting standard uncertainty when these two contributions are added
in square amounts to:
u( ax*' )=u( b x*' )=u( a*' *'

y )=u( b y ) = u 2p + ucc
2
= u2p + u((ekl)x )2
Anticipating the standard is approximately aligned with its rows and col-
umns along the x- and y-axes, this is estimated to be a function of scan
size and pitch.
For calibrations using the 895 nm reference standard (see section 2.4) at
scan areas of 50µm × 50µm to 70µm × 70µm we estimate:
u( ax*' ) = u( b x*' ) = u( a*' *'

y ) = u( b y ) = 0.22 nm
3.5.4 Environment: The mechanical stability δxM and δyM

The corrections δxM and δyM (for the x and y direction respectively) is a
sum of
• the change in the dimensions of the material connecting the sensing
tip and the distance sensor relative to the scanning area.
• the change in the dimensions of the material connecting the sensing
tip and sample relative to the scanning area.
It is changes in the environment temperature and changes in the internal
dissipation in the microscope which cause the material to expand or con-
tract during the image recording. The thermal contractions and expansions
cause the tip to be observed to drift over the surface when it is supposed
to stand still. The average position change is estimated to be zero and the
long term average drift speed is estimated to be 0 nm/s. Hence, δxM = δyM =
0.
However, during the time it takes to record one image a finite drift can be
observed. We interpret this as the uncertainty on δxM and δyM. From hundreds
of measurements we estimate average drift speeds (vdx and vdy) in x and y
during the time of the recording of one image to:
vdx = vdy = 0.2 nm/s during initial operation of the microscope.
vdx = vdy = 0.05 nm/s after 1 hour of scanning in a stable environment.
The terms δxM and δyM are corrections to the recorded pixel positions and
quantify the total drift in x and y, respectively, during the scanning of
one image, relative to the respective side lengths, S x* and S *y ,of the im-
age. Taking these numbers as the standard uncertainties of δxM and δyM we
get:
v dx(m / fs ) v dy(m / fs )
u(δ xM ) = , u(δ yM ) =
S x* S *y
where m is the number of scanned lines in the image fs [Hz] is the rate at
which lines are recorded. Thus 1/fs is the time it takes the microscope to
scan one line forward and one line back (trace and retrace).
For images with nominally equal side lengths in the range of 50µm × 50µm to
70µm × 70µm, with 512 lines, and for scan rates of 0.10-0.15Hz, after one
hour of scanning we estimate:
u(δxM) = u(δyM) = 0.005
3.5.5 Environment: Stability of capacitive sensors δxC and

δyC.
The correction δxC and δyC (for the x and y direction respectively) are the
relative changes in sensitivity of the capacitive sensors due to changes in
environmental parameters and electromagnetic interference around the ca-
pacitive plates in the distance sensors. The correction terms are estimated
to be zero, that is,
δxC = δyC = 0 .
The uncertainty is discussed in the following.
The capacitance of an air gap capacitor is proportional to the (relative)
permittivity, εr, of the air in the gap. An analysis of the influence of
changes in temperature, pressure, CO2 content and humidity of the air
within normal limits (much larger than those specified for our laboratory)
shows that the influence is less than 0.05‰. This is much smaller than
other uncertainty contributions, and can therefore be ignored. However, it
is known that amplifier drift and water condensation on the capacitor
plates lead to much higher changes in capacitance [2] than this. This
agrees with our results from more than twenty calibrations where a standard
deviation of the results of 0.3-0.4‰, which can be attributed to the dis-
tance sensors, has been observed. It is thus estimated that uncertainties
in the indication of the distance sensors lead to uncertainties of the cor-
rection terms δxC and δyC of:
u(δxC) = u(δyC) = 0.0004
3.5.6 Microscope reference frame (Abbé offset), δxA and δyA

The correction δxA and δyA (for the x and y direction respectively) are the
relative change in sensitivity due to the Abbé offset, caused by tilting of
the probe as it moves laterally. It is estimated to be zero, that is,
δxA = δyA = 0 .
Correcting the Abbé offset error requires precise knowledge of both the
tilt angle and probe height.
Based on the approximate length of the scanning device 50 mm and the ob-
served difference in probe height of approximately 50 µm, the relative Abbé
error is judged to be much smaller than (50 µm)/(50 mm) = 0.001 and at a
maximum 0.00005, that is
u(δxA) = u(δyA) = 0.00005
3.6 Uncertainty budget

An example of an uncertainty budget for the calculation of cxx* using DFM
GUM (the correctness of all uncertainty budgets have also been evaluated
manually):
DFM-GUM ver. 2.1a
Uncertainty Budget: Cx (60µm x 60µm image, after scanning one hour)

i Quantity (unit) Distribution xi u (x i ) νi ci u i (y ) r (x i ,y )
1 Reference length, La [nm] Normal 895 0.5 infinity 0.0011178 5.59E-04 0.764
2 Reference length, Lb [nm] Normal 895 0.5 infinity 1.831E-07 9.15E-08 0.000
3 Angle between La and Lb, γ [°] Normal 91.106 0.033 infinity 0.0001113 3.67E-06 0.005
4 Unit cell coordinate, a*'x [nm] Normal 894.38 0.22 infinity -0.001119 -2.46E-04 -0.336
5 Unit cell coordinate, a*'y [nm] Normal 5.8024 0.22 infinity -8.29E-06 -1.82E-06 -0.002
6 Unit cell coordinate, b*'x [nm] Normal -29.705 0.22 infinity 7.134E-06 1.57E-06 0.002
7 Unit cell coordinate, b*'y [nm] Normal 909.81 0.22 infinity 5.284E-08 1.16E-08 0.000
8 Correction for mechanical drift, δxM Normal 0 0.005 infinity 0.0019543 9.77E-06 0.013
9 Correction for capacitve sensor stability, δxC Normal 0 0.0004 infinity 1.000625 4.00E-04 0.547
10 Correction for Abbé offset, δxA Normal 0 0.00005 infinity 1.0006248 5.00E-05 0.068
11 Number of scanned lines in y , m Normal 512 0 infinity
Cx Normal 1.000624794 0.000732 infinity
Conf. level = 95.45% k= 2.0000

Result = 1.0006 U= 0.0015
Model: Y =-(SQRT((X7^2)*(X1^2)+(X5^2)*(X2^2)-2*X5*X7*X1*X2*COS(RADIANS(X3))))/((X5*X6-X4*X7)*(1-X8/X11-X9-X10))
The results for all three correction parameters are summarised in Table 2.
The uncertainties corresponds to the combined standard uncertainties multi-
plied by the coverage factor k = 2, in accordance with EAL-R2 [15].
Table 2. Example of calibration uncertainty (60µm × 60µm image)
Correction parameter Estimated value Uncertainty U

Cxx* 1.0006 0.0015
Cyy* 0.9834 0.0099
Cxy* 0.009479 0.0010
4. Vertical calibration using transfer standards

A thorough analysis describes the physical z-coordinate of an imaged sur-
face as a function of the observed and uncorrected z-coordinate and the
horizontal position. The two most important correction terms in a Taylor
expansion of this function are identified and assessed. A scaling factor in
the z-direction is calibrated based on series of measurements on a cali-
brated step height. By the use of a flat reference surface it is controlled
that the image bow is within limits.
4.1 Theory for vertical calibration

The definition of a step height is given in Figure 7.
The observed and uncorrected z-coordinate z*(x*i, y*j) as function of the
lateral position is given in equation (1) and discussed further in section
3.1.1. A single line (j = 1) is named a profile z*(x*); the average profile
to be used will be defined exactly in section 4.1.2.3.
4.1.1 The correction parameters czz* , czz*2 , and czx*2

We want to find a transformation of the (displayed) and uncorrected z* pro-
file z*(x*) into a metric coordinate system, that is z(z*(x*)) ≅ z(z*(x)) .
Contrary to the lateral plane this transformation cannot be approximated by
a linear function.
The metrology AFM system, including the readings from the position sensors,
can be described as having no significant hysteresis or time dependent
creep. Capacitive sensors does not suffer from these artefacts, and the me-
chanical part will not contribute significant. This has been verified for a
microscope similar to the one used here in [16]. On a timescale larger than
0.5 second, the response of the capacitive sensors and the simultaneous re-
sponse from an optical interferometer reading were compared. The difference
was observed to settle to a difference smaller than 1 nm one second after a
0.5 µm step was applied. The correctness of this basic assumption can be
assessed on a timescale of ten second or less by comparing the difference
between the profiles of a trace and a retrace of the tip following the same
line on the surface.
This implies that for each pair of uncorrected coordinate z* and position
x* in a profile there is, to a very good approximation, one and only one
position of the tip. This means that the metric z-coordinate z is a single
valued [17] function of the uncorrected z-coordinate z* and the uncorrected
position x* and thus the metric z-coordinate can be expressed as z(z*(x*)).
Let x*l be the scan length in the x-direction and let xi* = x*i – x*l/2 be
renamed x*i . The metric z-coordinate z(z*(x*)) can now be approximated by
a Taylor series [18] to second order in z*(x*) and x*, that is,
z(z* (x* )) = C00zz * + Czz *z* (x* ) + Czx *x* +

(27)
* * 2 *2 * * *
Czz *2 z (x ) + Czx *2 x + Czz * xz (x )x + L
This approximation will fail if the physical (that is the “true”) z-

coordinate z is not a single valued function [9] of the uncorrected z-
coordinate z*(x*) and the position x* .
The term C00xx* can be ignored as an irrelevant offset.
15
5
z [nm]
-5
-15
0 20 40 60
x [µm]
Figure 7 The top is the definition of a step height in analogy to the ISO stan-
dard 5436. The bottom is the interpretation of the definition for an average
profile. The solid lines is the three parts of two parallel lines fitted to the
profile in the three segment A, B and C .
The auxiliary z-coordinate z(A ) , z(B ) and z(C ) is defined as the average z*-
coordinate of the segment A, B and C of the profile (see Figure 7). The av-
erage off-set z*offset for the profile segment is then defined as
 z(C) + 1
z(A) + 1
z(B)
z*offset =  2 2 

(28)
 2 
The reference offset z*offset,ref is an average offset z*offset with an absolute
value of less than 500 nm. Let ∆h be a calibrated step height and ∆z* the
uncorrected difference in z*-coordinate values for the top and bottom line.
The coefficient
∂z ∆h
Czz * = ≅ (29)
∂z * z* = 0 ∆z * z *offset = 0
is the linear correction factor and is estimated as the correction factor

for a calibrated step height at offset zero.
The coefficient
∂z
C zx * = (30)
∂x * x* = 0
is zero as the X axis and the X* axis are chosen to be parallel and as the
Z axis is perpendicular to the X axis.
The coefficient
∂ 2z
c zz* 2 = (31)
2∂z*2 z*offset=0
is a measure for the linear variation in the linear correction factor as
function of the average off-set. It is estimated as the slope of the cor-
rection factor Czz* as function of the average z-offset, similar to the pro-
cedure in [16].
The coefficient
∂ 2z
Czx *2 = (32)
2∂x*2 x* = 0
is a measure for the image bow. It is estimated as minus the second order
coefficient in a polynomial fit to the average profile z’(xi) of a horizon-
tal and flat surface. To get a more useful and intuitive scale, this cor-
rection factor is quantified as the minimum to maximum value εp of the fit-
ted second order polynomial, over the maximal scan length xlmax , that is
2
x 
ε p = Czx*2  l max  (33)
 2 
The term Czx*2 z*(x)x can be ignored as the observed step height is found
not to depend on the horizontal position. The possible significant contri-
bution from higher order terms will be assessed in the uncertainty budget.
If the Z* axis is perpendicular to the X*Y* plane and the microscope coor-
dinate system was a linear function of the position then czz* would be a
scaling factor in the z-direction and all other coefficients would be zero.
The correction coefficients czz* , … are functions of the same parameters as
the sampled coordinates (see section 3.1.1). The measurand during calibra-
tion is defined as the values at equilibrium, at the reference temperature
Tref, the reference relative humidity Href, and a reference off-set
z*offset,ref , that is,
czz * ≡ czz *(Tref, H ref, z*offset,ref)
( 34 )
czz *2 ≡ czz *2(Tref, H ref, z*offset,ref)
czx *2 ≡ czx *2(Tref, H ref, z*offset,ref)
For further details about influence parameters see section 4.5.
4.1.2 Plane correction

The thermal drift in the z direction during the scanning implies that the
AFM does not really deliver a topography image, but only neighbouring pro-
files, whose relative position are not known exactly as discussed in sec-
tion 3.1.4.
4.1.2.1 The image bow

To remove the image bow from the image of the step height (due to the cor-
rection term Czx*2) an artificial image with the opposite bow is subtracted.
On a “Profile” of the artificial image it is controlled that a second order
“Curve fit” gives a coefficient to the x2 term equal to minus the correc-
tion coefficient Czx*2 given in the registrations for the instruments (“ap-
paratur kort”).
4.1.2.2 Line wise levelling

For an exact mathematical description of the algorithms estimating the ref-
erence lines see Addendum (section 6).
For the images of the flat reference surface “Line wise levelling” is done
once using a 1. order polynomia fit in the “Plane Correction” Menu under
“Processing” in SPIP.
For the images of step heights “Line wise levelling” is performed itera-
tively 3-4 times using 1. order polynomia, restricting the estimation vol-
ume to the basal plane2 of the image using the “Inside Color Range” option
in the “Plane Correction” Menu.
4.1.2.3 The average profile z*(x*)

To simplify the mathematics, the images are treated as m profiles along the
fast scanning x direction, and the calibration is done (and is only valid)
for a profile or an average of profiles. An average profile is calculated
as follow
m
1
z**(x *i) =
n
∑ z*(x*i, y *j) (35)
j=1
where z*(x*i, y*i) is defined in equation (11). To simplify the notation

z**(x*) is renamed z*(x*).
For the image of the step height and the flat reference surface an average
profile is calculated using “Average X-Profile” in “Averaging” in the
“Processing” menu.
4.1.2.4 Correction for the surface tilt

The software maintains the original x* and y* scale after levelling, see
the discussion in section 3.1.4. To correct for this the microscope coordi-
nate z* should be transformed as follow
z* → z* cos β x cos β y (36)
z' → z' cos β x cos β y (37)
where βx and βy are the tilt angles of the surface of the standard with re-
spect to the scanning plane along the x-axis and the y-axis, respectively.
In practice the tilt angles, βx and βy, are often below 0.5°, which implies
the cosines to deviate from unity with less than 8×10-5. Therefore the
cos(βx) and cos(βy)terms are considered unity in the analysis and the asso-
ciated uncertainty vanishing. For the y-axis βy is estimated as the average
of the up and down scan, for the x-axis βx is estimated as the average of a
trace and retrace scan. If it is higher than 0.5° the images are corrected
by multiplying the z’ with cos(βx)cos(βy) before the plane correction is ap-
plied
2
4.1.3 Determination of the observed step height

The observed height ∆ho , that is, the difference in z-coordinate for the
top and bottom part for an average profile is fitted from
 
 
min ∑ (z* (xi ) − ho ) 2 + ∑ z* (xi ) 2  (38)
ho
 x *i ∈ C x *i ∈ A 
 x *i ∈ B 
where the profile segment A, B and C are defined in Figure 7, The step
height is calculated using “ISO 5436” under “Z-calibration” under “Process-
ing”. In the windows, which give the parameters for the calculation “Middle
Length”, “Side Length” and “Edge Distance” should all be set to 33.00.
“Groove” or “Line” should be checked as appropriated. The button “Profile”
should be marked.
4.1.4 Estimation of correction parameters czz* , czz*2 , and

czx*2
4.1.4.1 Estimation of czx*2

For six average profiles of the flat reference (from 60 µm × 60 µm images
evenly distributed over the certified area) the second order coefficient to
the x2 term using “Curve fit” is recorded. The average and standard devia-
tion is calculated.
4.1.4.2 Estimation of czz*2

For approximately 6 different average profiles (from images 60 µm × 60 µm
corrected for image bow) of the reference standard with z offset in range
between approximately ± 2 µm, consecutive values of the average z-offset
(see equation (28)) and the observed height is recorded. The correction pa-
rameter czz* is then calculated and transferred to an excel spreadsheet.
From a linear fit to czz* as function of the offset in micrometer the linear
coefficient czz*2 is calculated in the unit 1/µm (see checklist and Figure
8).
y = -0.0018x + 1.0019
1.006
R2 = 0.9584
1.004
1.002
H800
1.000
H80
0.998 Linear (H800)
Linear (H80)
0.996
0.994
0.992
-3 -2 -1 0 1 2 3 4
Figure 8 Example of the measured correction factor czz* as function of the average
observed z-coordinate. The squares are correction factors for ca step height of
nominal 800 nm; the diamonds are correction factors for a nominal step height of 80
nm. The non-linear variation of the correction factors as function of the average
z-position is insignificant.
The size of czz*2 is compared with the statement about the nonlinearity of
the z-sensitivity in the registrations for the instruments (“apparatur
kort”) and based on the measurements it is assessed if this statement is
still valid. If the value is significant different from the value in the
registrations for the instruments proper action is taken and documented on
the registration for the instrument or on the internal certificate.
4.1.4.3 Estimation of czz*

Let href be the reference height of the nominal 800 nm step height and
hobs,ref be the observed height of the reference. The correction factor czz* ,
is then
href
czz * = (39)
hobs,ref
4.2 Measurement of reference standard

The default scan angle is 0°, see the discussion in section XX.
Before acquiring the first calibration images, after the window enclosing
the microscope has been opened, the microscope should have been scanning
continuously for one hour or more. For some images, this is fulfilled by
using only the second image for calibration. The filenames is recorded
Two images are recorded (trace, retrace and amplitude), however, it is the
“Height” information in the second image with direction “trace” and “down”,
which should be used for the calibration. The minimum and maximum tempera-
ture and humidity under the calibration is recorded. In order to calibrate
the instrument in its full measurement range the largest possible quadratic
scan area is chosen. This will usually be around 60µm × 60µm.
4.2.1 Measurement of czx*2

The parameter czx*2 is found by acquiring the images according to section
4.1.4.1 and Table 3.
Table 3 Parameters for imaging the flat reference

tion rate direction
Flat refer- 60µm × 60µm 512 × 64 0 0.5 Hz Trace up

ence
Flat refer- 60µm × 60µm 512 × 64 0 0.5 Hz Trace down
ence
4.2.2 Measurement of czz*2

The parameter czz*2 is found by acquiring the images according to section
Table 4 Parameters for imaging the step height for measuring czz*2

tion rate direction
Standard 60µm × 60µm 512 × 32 0 0.14Hz Trace up

H800
Standard 60µm × 60µm 512 × 32 0 0.14Hz Trace down
H800
4.2.3 Measurement of czz*

The parameter czz* is found by acquiring the images according to section
Table 5. Parameters for imaging the step height for measuring czz*2

tion rate direction
Standard 60µm × 60µm 512 × 512 0° 0.14Hz Trace up

H800
Standard 60µm × 60µm 512 × 512 0° 0.14Hz Trace down
H800
4.3 Image processing and calculations

For the estimation of czx*2 and czz*2 the results are typed into two excel
sheet and saved as <imagefilename>.xls .
The model function to calculate czz* , czx*2 and czz*2 is typed into DFM-GUM
templates. To rule that the typing was done correct, it is controlled that
appropriate values in the uncertainty budget give the same results. Then
the observed height of the reference step height is typed into DFM-GUM
worksheets and the calculated correction parameters and uncertainties are

recorded. The worksheets are saved as <imagefilename>_Cx.xls etc.
4.4 The certificate

Before the certificate is written the calibration should be accepted: The
parameters czx*2 and czz*2 should be ≈ 0, and czz* ≈ 1. Additionally the dif-
ferences to previous values should be reasonable.
4.5 Uncertainty analysis

The input quantities to the model function are significantly influenced by:
• The accuracy of the calibration of the reference standard.
• Lack of accuracy when recording and processing the image.
• The lack of equilibrium and knowledge about the environmental condi-
tions.
• The lack of perfect movement of the probe in the microscope coordi-
nate system.
Other influence sources have been considered and found not to be signifi-
cant. These are, the horizontal orientation of the grating, the tip shape,
a small amount of dust on the sample, electromagnetic interference from the
power line, acoustical noise and the force exerted on the sample by the
tip.
The evaluation of the standard uncertainty on the input quantities is based
on the experience and experimental documentation in the laboratory summa-
rized in the scientific publications [18], the Ph.D. thesis [12] and the
report on the international supplementary key comparison NANO2 (results
will be published).
4.5.1 The model function

The auxiliary z-coordinate z(A ) , z(B ) and z(C ) is defined as the average z*-
coordinate of the segment A, B and C of the profile (see Figure 7); they
are defined in a similar way for a profile of a flat surface. The differ-
ence in z-coordinate h’obs,ref for the profile segment A, B and C is then
 (z(A ) + z(B )) 
h'obs,ref = z(C ) −  ≈ hobs,ref
 (40)
 2 
Let aref x*2 be the reference bow of the flatness standard, that is, the sec-
ond order coefficient in a polynimial fit to the average profile, see equa-
tion (32), let ax *2 be the observed average curvature and let t be the time
it takes to scan an image. The corrections parameters are given by
czx *2 = ax *2 + aref x *2 + δzt*2t2
czz*2 = czz*2 (41)

href
czz* =
hobs, ref(1 + δzz*)
where the corrections δzt*2 and δzz* are defined in the following sections.
4.5.2 The reference standard: The certified height href

The estimate is the dimensions given in the certificate (h ≈ 760 nm). The
standard is manufactured from silicon and silicon oxide for which the ther-
mal expansion coefficient at room temperature is about 5×10-6 K-1 and 5×10-
6
K-1 which is insignificant compared to the other larger contributions to
the resulting uncertainty. Temporal drift of silicon is in the order of 10-8
per year, which is also insignificant. Thus the uncertainty is estimated to
be the uncertainty on the calibration stated in the certificate:
u(href) ≈ 4.1 nm
4.5.3 The reference standard : The flatness reference

aref x*2
Based on measurement by PTB on similar samples the image bow is estimated
to zero and the uncertainty is estimated to (for a side length of 65 µm)
u(aref x *2 ) = 4.7 ⋅ 10−10 nm-1
4.5.4 Profile quality and processing: The observed step

height hobs,ref and image bow ax *2
The estimates are calculated by [10] based on a tilt corrected profile. The
calculation by [10] and the tilt correction of the profile introduce some
errors due to:
• the limited number of pixels, and recorded lines
• the limited accuracy when removing the tilt and offset by the parameters
a, and b
• imperfections in horizontal alignment of the grooves in the image before
the average profile is calculated (only for step heights)
• imperfections in estimation of the edge position and there by the A, B
and C segment of the average profile (only for step heights)
• the difference between the calculated projected step height or image bow
for the observed profiles and the step height or image bow perpendicular
to the surface.
It is estimated to be
u(href,obs) = 0.0002*href,obs + 0,02 nm
u(aref x *2) = 0.001 nm
4.5.5 Environmental: The mechanical stability δzt*2

δczt*2 is the (random) change of z (A ) and z (B ) relative to z (C ) due to
(mostly thermal) drift and is estimated to be zero. The uncertainty u(δczt*2)
will be smaller as the number of uncorrelated lines and images increase. It
is estimated to be (for a side length of 65 µm)
u(δzt*2t2) = 9.5 ⋅ 10-11 nm-1 (128 lines or more)
u(δzt*2t2) = 1.9 ⋅ 10-11 nm-1 (64 lines or less)
4.5.6 Environmental: Stability of capacitive sensors δzz*

The correction δczz* is the relative change in sensitivity of the capacitive
sensors due to changes in the environmental parameters, see the discussion
in section 3.5.5. It is estimated to be zero with a standard uncertainty of
u(δczz*) = 0.0004
4.5.7 Microscope reference frame (nonlinearity) czz*2

The correction czz*2 is minus the error due to remaining nonlinearity of the
z-scale of 2.order or higher in z*. This is because the compensation done
by the correction factor czz*2 is not complete. It is estimated to be zero
with a standard uncertainty of
u( czz*2 ) = 0.00002
5. References to literature
[1] Das Metrologie-Rasterkraftmicroskop, F. Meli, OFMET info, Vol. 6

no. 1, pp1-8, Switzerland (1999)
[2] The Nanopositioning Book, T. Hicks, P. Atherton, Queensgate Instru-
ments Ltd. ISBN 0 9530658 0 4(1997).
[3] The validation of the software is carried out according to the RL10
guidelines by the Danish accreditation body DANAK.
[4] National Physical Laboratory (NPL), United Kingdom by the use of
Littrow diffraction (green He-Ne laser) using a manual angle table
with two readings on an optical screen.
[5] Made by Nanosensors see also J. Garnaes, N. Kofod, J. F. Jørgensen,
A. Kühle, P. Besmens, O. Ohlsson, J. B. Rasmussen, P. E. Lindelof,
G. Wilkening, L. Koenders, W. Mirande, K. Hasche, J. Haycocks, J.
Nunn, M. Stedman, Nanometre scale transfer standards, Proceedings
for euspen 1st international conference and general meeting of the
european society for precision engineering and nanotechnology, Ed-
ited by: P. McKeown, J. Corbett et al., on May 31st - June 4th 1999
Congress Centre Bremen, Germany, Vol 2, 134-137 (1999)
[6] Physikalische-Techische Bundesanstalt (PTB) certificate 146 PTB 99
[7] Dimension™ 3100 with metrology head from Digital Instruments (now
Veeco), Santa Barbara, CA, USA, www.di.com.
[8] Methods for lateral calibration of Scanning Probe Microscopes based
on two dimensional transfer standards, N. Kofod, J. Garnaes, J. F.
Jørgensen, Proceedings of the 4th seminar on Quantitative Microscopy
QM 2000 Dimensional measurements in the micro- and nanometre range,
Edited by Klaus Hasche, Werner Mirandé, Günter Wilkening, Semmer-
ing, Austria, January 12-14 2000, PTB-Bericht, page 36-43 (2000)
[9] Calibrated line measurements with an atomic force microscope,
N. Kofod, J. Garnaes, J. F. Jørgensen, European society for preci-
sion engineering and nanotechnology: Proceedings for the 1st Topical
Conference on Fabrication and Metrology in Nanotechnology, Edited
by L. De Chiffre, K. Carneiro, Copenhagen May 28-30, Vol. 2, page
373-381 (2000)
[10] Scanning Probe Image Processor, Image Metrology Aps., Nils Koppels
Allé 402, DK-2800 Lyngby, Denmark. www.imagemet.com.
[11] Scanning Probe Microscopy Image Restoration and Analysis, J.Friis
Jørgensen, Ph. D. Thesis, Technical University of Denmark 1993 ISBN
87-88306.
[12] Validation and industrial application of AFM, Niels Kofod, Ph.D.
thesis, IPL Technical University of Denmark, IPL.004.02. (2002).
[13] Manuals for the Dimension™ 3100, and Support Note No. 242 Rev. B.
Metrology scanning probe microscopy – description and use. See [7].
[14] WGDM-7: Preliminary comparison on nanometrology according to the
rules of CCL Key Comparisons; NANO4: 1D Gratings Final Report Draft
B. Edited by F. Meli, Suisse Federal Office of Metrology, November
2000. Included in the BIPM key comparison database.
[15] ISO/BIPM,”ISO Guide to Expression of Uncertainty in Measurement”,
Corrected and reprinted, 1995, 1993(E)
[16] Z-calibration of a metrology AFM scanner using an interferometer

and a tilting device together with a linear displacement stage F.
Meli, Proceedings of the 3rd Seminar on Quantitative Microscopy,
November 1998, Lyngby, Denmark, PTB-F-34 p61-67 (1998)
[17] A “single valued function” is normally just referred to as a “func-
tion” but the phrase “single valued” is used here to point out the
difference to a “multi valued function”. For a piezo electrical
scanning system exhibiting hysteresis and creep the position of the
tip as function of the control signal (typical the voltage applied
to the peizo scanner) is a multi valued function, as there are sev-
eral possible positions to a given control signal depending on the
history of the control signal applied and the time since the appli-
cation of the voltage started (creep).
[18] Calibration of step heights and roughness measurements with atomic
force microscopes J. Garnaes, N. Kofod, A. Kühle, C. Nielsen,
K. Dirscherl, L. Blunt, accepted for publication in Precision Engi-
neering DFM-01-R40 (2001)
6. Addendum - Estimation of a reference plane, levelling

Let ηj (xi ) = a0j + a1jxi + a2jxi 2 +... define a z' (xi , y j ) ,
line. Let
i = 1,...n,j = 1,...m , be the observed height of the surface. A reference plane
η(xi, y j) is then estimated based on the observed lines z'j (xi ) by subtract-
ing a reference line ηj(xi ) estimated by the least square method. The coef-
ficient A 0j , A1j , A 2j ,... ∈ {a0j , a1j , a2j ,...} to the reference line ηj(xi ) is then es-
timated based on the observed line z'j (xi) by minimising ε j2 with respect to
a0j, a1j, a2j,... in the following equation:
n
ε j2 = ∑ (z'j (xi ) − (a0j + a1jxi + a2jxi 2 +...)) 2 (42)
i =1
Restricting the estimation volume to the basal plane3 of the image using
the “Inside Color Range” option in SPIP means that only observed heights
z'j (xi ) in the basal plane is included in the minimising.
The reference plane is then given by
η (xi , y j ) = A 0j + A1jxi + A 2jxi 2 +... .
(43)
A corrected image z(xi, y j) is then calculated by subtracting the reference
plane from the observed height of the surface
z(xi, y j) = z'(xi, y j) − η(xi, y j)
(44)
If the highest coefficient for x is 1, that is, η j(xi ) = a0j + a1jxi the ref-
erence line is named a 1. order line estimated based on the least square
method. If the highest coefficient for x is 3, that is,
2 3
η j(xi ) = a0j + a1jxi + a2jxi +a3jxi the reference line is named a 3. order line
estimated based on the least square method.
3

NT TR 536 Calibration Procedures For Atomic Force Microscopes (Nordtest Technical Report)

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Nordtest Technical Report 536

Calibration Procedures for Atomic Force Microscopes

Figure 1. Principle of the atomic force microscope.

1.2 Project background, objectives and main results

1.4 Course of the project

2 Writing of detailed calibration procedures DFM

3 Test and evaluation of procedures SP, DFM

4 Test on standard AFM (non metrology grade) SP

5 Comparative measurements of test specimens TPN, SP, DFM

Figure 2. Initial project schedule. X = Milestones.

Standard Ibsen 2D1000, S/N A00051

2.1 Calibration of the metrology AFM

2.2 Calibration of Toponova’s standard AFM

2.3 Calibration of SP’s standard AFM.

2.4 Calibration of Toponova’s interference microscope.

Standard Nanosensors H800, SN

href (759.7 ± 8.2) nm

3.1 Calibration of the metrology AFM and of the TGZ02 grating.

3.2 Calibration of Toponova’s and SP’s standard AFMs

3.3 Measurement of the TGZ02 grating with an interference microscope.

4 Comparative roughness measurements on ball-joints for hip implants

4.1 AFM roughness measurements

Partner Hip joint 1DP Hip joint 2BM

Hip joint ball 1DP Hip joint ball 2BM

4.2 Comparison between AFM and interference microscope roughness data

Partner Hip joint 1DP Hip joint 2BM

6 List of equipment used

Nordtest Technical Report 536

Procedures for the calibration of the three axes of an

atomic force microscope by the use of transfer standards

1.1.1 The metric coordinate system

2.1 The microscope

2.2 Clean room bench

2.4 The two-dimensional standard

Figure 2. Drawing of the

2.5 The step height standard

film metallic film (PtIr-Cr) approximately 70 nm in thickness, see Figure

3. Lateral calibration using transfer standards

3.1 Theory for lateral calibration

3.1.1 The microscopes coordinate system

3.1.2 The actual traced coordinate system

3.1.3 The correction parameters cxx*, cyy*, and cxy*

For further details about influence parameters see section 3.5.

3.1.4 Plane correction

 x*'  x*' / cos βx 

3.1.5 Determination of observed unit cell features

2. The cross-correlation function CAu(x*,y*) between the particular unit

C Au(x*, y*) = ∑ z(x* + c* xo, y* + c* y p) ⋅ zAu(c* xo, c* y p) (6)

{p* kl} = {(x*, y*) C Au(x*, y*)(x*,y*) is a local maximum} (7)

where, k = 1, … K and l = 1, … L are indices for the cross-correlation

e kl = p* kl − (p* 00+ ka*+ lb*) (8)

where p*00 is the coordinates of the highest peak in the cross-

Recorded image Unit cell template Cross-correlation function

3.1.6 Estimation of correction parameters cxx*, cyy* and cxy*

3.2 Measurement of reference standard

3.2.1 Measurement procedure

Figure 6. The reference stan-

The microscope is operated in Tapping Mode according to the guidelines and

Table 1. Parameters for images used for calibration.

Standard Scan size Sampling Scan rota- Scan Scan

3.3 Image processing

is controlled that the appropriate values in the uncertainty budget in sec-

3.4 The certificate

3.1.3 The correction parameters cxx, cyy, and cxy*

 x'  x' / cos βx 

2. The cross-correlation function CAu(x,y) between the particular unit

C Au(x, y) = ∑ z(x* + c* xo, y* + c* y p) ⋅ zAu(c* xo, c* y p) (6)

{p* kl} = {(x, y) C Au(x, y)(x,y) is a local maximum} (7)

e kl = p* kl − (p* 00+ ka+ lb) (8)

3.1.6 Estimation of correction parameters cxx, cyy and cxy*

 x'  1 + δ xC 0 1 + δ xA 0 1 + δ xM /m δ xM  x 

 x' 1 + δxC + δxA + δxM /m δxM  x 

b'x ⋅ b'y ⋅ L2a + a'x ⋅ a'y ⋅ L2b − B ⋅ La ⋅ Lb ⋅ cos(γ)

B = (a'y b'x +a'x b'y ) (26)

u( ax' )=u( b x' )=u( a' '

u( ax' ) = u( b x' ) = u( a' '

4.1.1 The correction parameters czz* , czz2 , and czx2

z(z* (x* )) = C00zz * + Czz z (x* ) + Czx x +

4.1.2.3 The average profile z(x)

where z(xi, y*i) is defined in equation (11). To simplify the notation

czx 2 = ax 2 + aref x 2 + δzt2t2

czz2 = czz2 (41)