on the surface. The cantilever should not be too stiff as it can
go too deep and damage the sample.
9. Place a label on the slide with a permanent marker to help to
focus on the slide surface.
10. If the fitting is not satisfying, change the Markers to 3, 5, or
7 to smooth the line and get the best reduction of the thin
individual peaks (spikes) and click again on fit data.
11. The measurement is done in the air on dry samples. Dust can
cause artifacts during the scan, so use the compressed air duster
to clean the stage and slide.
12. Localize the section by looking at the marker label below the
slide.
13. The AFM scan is time-consuming, and a detailed scan usually
takes a long time (depending on the number of pixels, usually
around an hour). Images recorded with decreased Samples/
Line help to save time before ROIs are found. Sometimes, the
artifacts are present on the images; if so, the scan should be
repeated.
14. Make sure that the resolution is good enough to record a
sufficient number of force curves within the ROIs.
15. The DMT model calculates the contact between the tip of
cantilever and the sample by measuring the load and adhesion
forces outside the contact area.
16. Before extracting the force curves, rotate the images in 3D to
see if the section is flat and if there are no artifacts on the
surface. Make sure that the group of analyzed cells form a flat
surface. First, select random pixels in different regions of the
image and check if the force curves corresponding to these
pixels (Fig. 4c) are correct (see model force curve Fig. 1b).
17. Each ROI should consist of at least 100 force curves; but the
more, the merrier. Always make sure the ROI has enough force
curves to perform statistical analysis.
18. The fixation and resin embedding modifies the properties of
the sample, but if performed at an organ scale it should preserve relative stiffness within the cell walls.
19. The R
2 should be more than 95%, the force curves with low
quadratic fit (<95%) are excluded. If more than 5% of the force
curves need to be excluded within the image, then the image is
discarded.
20. The AFM measurements are relative and depend on individual
samples. The percentage is based on the difference in stiffness
among different ROIs within the same sample.
Atomic Force Microscopy to Study Cell Wall Mechanics in Plants
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go too deep and damage the sample.
9. Place a label on the slide with a permanent marker to help to
focus on the slide surface.
10. If the fitting is not satisfying, change the Markers to 3, 5, or
7 to smooth the line and get the best reduction of the thin
individual peaks (spikes) and click again on fit data.
11. The measurement is done in the air on dry samples. Dust can
cause artifacts during the scan, so use the compressed air duster
to clean the stage and slide.
12. Localize the section by looking at the marker label below the
slide.
13. The AFM scan is time-consuming, and a detailed scan usually
takes a long time (depending on the number of pixels, usually
around an hour). Images recorded with decreased Samples/
Line help to save time before ROIs are found. Sometimes, the
artifacts are present on the images; if so, the scan should be
repeated.
14. Make sure that the resolution is good enough to record a
sufficient number of force curves within the ROIs.
15. The DMT model calculates the contact between the tip of
cantilever and the sample by measuring the load and adhesion
forces outside the contact area.
16. Before extracting the force curves, rotate the images in 3D to
see if the section is flat and if there are no artifacts on the
surface. Make sure that the group of analyzed cells form a flat
surface. First, select random pixels in different regions of the
image and check if the force curves corresponding to these
pixels (Fig. 4c) are correct (see model force curve Fig. 1b).
17. Each ROI should consist of at least 100 force curves; but the
more, the merrier. Always make sure the ROI has enough force
curves to perform statistical analysis.
18. The fixation and resin embedding modifies the properties of
the sample, but if performed at an organ scale it should preserve relative stiffness within the cell walls.
19. The R
2 should be more than 95%, the force curves with low
quadratic fit (<95%) are excluded. If more than 5% of the force
curves need to be excluded within the image, then the image is
discarded.
20. The AFM measurements are relative and depend on individual
samples. The percentage is based on the difference in stiffness
among different ROIs within the same sample.
Atomic Force Microscopy to Study Cell Wall Mechanics in Plants
367