NanoLocz

NanoLocz User Guide 1.3

Installation - - Getting Started - - Saving Data
Level - - Detect - - FineAlign - - Localize
Area Analysis - - Keyboard Shortcuts
Simulation AFM - - YouTube Video Tutorials

Image Levelling

Manual Leveling

Plane Leveling:
To remove tilt or bow in images:

• Set the x and y Plane values to perform polynomial fitting to all the x and/or y values in each image.
• Plane fitting is previewed in real-time.
• Press the ‘Accept’ button to apply.
• Tip: X and Y values of 1 will remove linear tilt, values of 2 or more can remove image bow.

Line Leveling:
To remove height variations between scan lines:

• Set the x and y Line values to perform polynomial fitting to each image line in x and/or y.
• Line fitting is previewed in real-time.
• Press the ‘Accept’ button to apply.
• Tip: use ‘-Med’ to remove the median of each line in x.

Line or Plane Leveling with a Mask:
To level images and movies that contain a variety of height levels requires certain regions (referred to as masked regions) to be excluded from the polynomial fits. Typically the included region should be a surface which is expected to be level and has a high coverage (>50%). The options to generate a mask are: 1) Using the upper and lower threshold values on the height histogram 2) Manually drawing using the Draw Mask button 3) Using the ‘Mask with z-scale’ to mask values outside of the colormap scale 4) Automatically using the Otsu method 5) Automatically using the Fit button (fits a Gaussian to all the values in the movie and using 1.5σ (standard deviation) as the threshold values.)

• To level with a mask, line leveling or plane leveling is then performed as before but fitting is performed using only the non-masked values.

• Press the ‘Accept’ button to apply.

  🛠 Automatic Leveling Routines

  NanoLocz provides several pre-configured automatic leveling routines to standardize and correct height data across AFM image stacks. These routines are tailored for different sample features.

plane-line

• Steps:
  • Subtracts a best-fit plane (x and y directions).
  • Applies a median line subtraction in x.

    multi-plane-edges

• Purpose: Edge-based masking for step detection -> useful if there are multiple flat surfaces
• Steps:
  • Plane subtraction.
  • Median line if vertical variation dominates.
  • Edge detection masking → used to apply weighted plane and line leveling multiple times.
  • Final mean-plane subtraction.

multi-plane-otsu

• Purpose: Uses Otsu method to generate edge masks for multi plane fitting -> useful if there are multiple flat surfaces
• Steps:
  • Plane subtraction.
  • Median line subtraction if vertical variance exceeds threshold.
  • Otsu edge masks applied iteratively for plane/line subtraction.
  • Final mean-plane subtraction.

iterative 1nm high, iterative -1nm low, iterative high low

• Purpose: Removes features above/below a fixed height.
• Steps:
  1. Plane subtraction.
  2. Line subtraction if vertical variance dominates.
  3. Iteratively thresholds using fixed z-limits and reapplies plane and median line subtraction with masking.

Line1 + Otsu Line2

• Steps:
  • Subtracts a linear line fit (first pass).
  • Applies Otsu thresholding to mask and subtracts a stronger 2nd order line fit.

high-low x2 (fit)

• Purpose: Uses Gaussian fitting on the histogram to determine leveling thresholds.
• Steps:
  • Initial plane + median line subtraction.
  • Histogram of z-values fitted with Gaussian.
  • Mask generated for values within ±1.5σ.
  • Leveling reapplied with mask.

iterative fit holes, iterative fit peaks

• Purpose:
  • Holes: For images with lower-intensity defects.
  • Peaks: For images with higher protrusions.
• Steps:
  1. 2nd-order plane fit + median line subtraction.
  2. Gaussian fit on z-values → ±1.5σ mask (shifted for holes or peaks).
  3. Repeat leveling with mask.
  4. Generate new mask and repeat.

  5. Final 1st-order line subtraction with mask.

Choosing the Right Routine

• Flat surfaces with sparse particles → iterative 1nm high or high-low x2 (fit)
• Surfaces with pits/defects → iterative fit holes
• Surfaces with sharp peaks → iterative fit peaks
• Samples with more than 1 plane → multi-plane-edges or multi-plane-otsu
• General leveling → plane-line or Line1 + Otsu Line2

YouTube video on Leveling + Area Analysis

This site is open source. Improve this page.