fwhm_on_spots

Full-width-half-maximum of 2D spots

This jython-fiji macro computes the FWHM on bright spots on dark background by, first, finding all maxima in an image (using fiji Find Maxima) and second, iterating over all 2D peak locations. For each peak the horizontal, vertical and two diagonal line intensity profiles are extracted and a general Gaussian function is fit. From that the standard deviation and goodness-of-fit is extracted. The standard deviation is converted to FWHM.

Peak locations, FWHM, goodness-of-fit and other read-outs are exported to an tab-delimited, excel readable file. In additions, all (fiji-)Rois of the according line profiles are exported into a fiji RoiManager zip-file.

Input

1. Image: A gray-valued image (already opened in fiji)
2. Peak tolerance: Height of the peak above background (see fiji Find Maxima)
3. Window radius for Gaussian fit in pixel
4. Pixel size in nano-meters (for export)
5. Output directory (for export)

Output

1. fwhm_values.txt: Columns are:

   • id: index
   • peak_id: number of peak
   • x_pos, y_pos: the x and y position in image
   • type: horizontal (H), vertical (V), diagonal 1 (D1), diagonal 2 (D2)
   • fwhm: full width half maximum of Gaussian fit
   • fwhm_nm: FWHM scaled by pixel size
   • r2_GoF: goodness of fit. Good values for a fit should be > 0.9
   • avg_fwhm: average of all fwhm of the four types (H, V, D1, D2) per peak_id
   • area_profile: area under the profile
   • area_gauss: area under the Gaussian fit

2. fwhm_fiji_rois.zip: Fiji-Rois with horizontal, vertical and diagonal line profiles named as "<index>_<peak_id>_<type>"

Installation

1. Install into fiji's plugin-folder

   Download and copy fwhm_on_spots.py to your the fiji plugin-folder

2. From scipt editor

   Download fwhm_on_spots.py and open it in fiji's script editor and run it from there

Example

You can download the used test image.