Puncta Analyzer

Quantify puncta in images

Requirements

• anaconda 3.7
• conda
• imageJ

Installation

Download this repository via git or github

Using:

• terminal for Mac OS
• conda prompt for Windows

navigate to repository on your computer after you download it e.g. and create the conda environment

cd ./puncta-analyzer
conda env create -f environment.yml

Usage

The puncta analyzer is a two step process

1. Image straightening
2. Image analysis

Image Straightening

1. in the dist folder, copy Preprocessor_2.jar to your imagej plugins folder
2. open the tiff file to analyze in imagej
3. use the segmented line tool to draw a series of line segments representing the region of the image you are interested in
4. open the plugins -> Preprocessor_2 plugin
5. save the straightened images to a directory of your choice
6. two images will be saved

• the brightened image is for presentations
• the processed images is for analysis
• generally, save the processed images in one directory so they can be batch processed by the puncta analyzer

Image Analysis

To run the program:

conda activate puncta-analyzer

Navigate to the directory that you downloaded puncta-analyzer to:

cd ./puncta-analyzer

Basic Usage

python src/puncta-analyzer.py <src> <dst>

Example:

python src/puncta-analyzer.py ./data/subtracted.tif ./data/

Options

To view all options, type:

python src/puncta-analyzer --help

Usage: puncta_analyzer.py [OPTIONS] INPUT_PATH OUTPUT_PATH

  analyze puncta in tif files

Options:
  -lo, --min-sigma INTEGER RANGE  Lower values detect smaller puncta
                                  [default: 1]
  -hi, --max-sigma INTEGER RANGE  Higher values detect larger puncta
                                  [default: 10]
  -t, --threshold FLOAT RANGE     Higher values remove puncta with duller
                                  intensity  [default: 0.1]
  -ov, --overlap FLOAT RANGE      If the fraction of overlap between two
                                  puncta is greater than this value, remove
                                  the smaller puncta  [default: 0.7]
  --save-annotated                Draw circles around recognized puncta and
                                  save as an image  [default: False]
  --help                          Show this message and exit.

Batch Processing

If INPUT_PATH is a directory, all tif files in the INPUT_PATH will be processed

Notes

• Make sure only tif files created from the Preprocessor_2 plugin are in this directory
• The program won't work for general tif files

Output

Analyzed stats and annotated images will be saved in the OUTPUT_PATH

Examples

if INPUT_PATH is a.tif Then OUTPUT_PATH will contain a-puncta-stats-<timestamp>.csv and puncta-img-set-stats-<timestamp>.csv

• a-puncta-stats-<timestamp>.csv will contain puncta specific stats
• puncta-img-set-stats-<timestamp>.csv will contain image specific stats