The GDSC Single Molecule Light Microscopy (SMLM) plugins provide various tools for single molecule localisation analysis. This includes PALM, STORM and other single molecule microscopy methods.

The plugins provide tools to:

• Fit an image, or series of images, using a 2D Guassian Point Spread Function (PSF)
• Save results to a table, a file, an image and/or to memory
• Trace or cluster localisations through time to identify molecules
• Correct drift in long time course images
• Estimate fluorophore dark-time and blinking rate
• Create localisation density images
• Create custom PSFs from calibration images
• Create simulated data using a Gaussian or custom PSF with configurable molecule populations and diffusion
• Calibrate the gain and read noise of your microscope camera
• Estimate noise in an image
• Estimate resolution using Fourier Ring Correlation

Results can be loaded from file for analysis using the following formats:

• SMLM Text/Binary
• RapidSTORM
• Nikon DSTORM

See the latest documentation on ReadTheDocs.

The SMLM plugins are distributed using an ImageJ2/Fiji update site.

To install the plugins using Fiji (an ImageJ distribution) just follow the instructions How_to_follow_a_3rd_party_update_site and add the GDSC SMLM update site. All the plugins will appear under the 'Plugins > GDSC SMLM' menu.

The source code is accessed using git and built using Maven.

The code depends on the gdsc-analytics, gdsc-test and gdsc-core artifacts so you will have to install these to your local Maven repository before building:

1. Clone the required repositories

    git clone https://github.com/aherbert/gdsc-analytics.git
    git clone https://github.com/aherbert/gdsc-test.git
    git clone https://github.com/aherbert/gdsc-core.git
    git clone https://github.com/aherbert/gdsc-smlm.git
   

2. Build the code and install using Maven

    cd gdsc-analytics
    mvn install
    cd ../gdsc-test
    mvn install
    cd ../gdsc-core
    mvn install
    cd ../gdsc-smlm
    mvn package
   

   This will produce a gdsc_smlm-[VERSION].jar file in the target directory. All dependencies are copied into the target/dependencies directory.

3. Copy the gdsc_smlm* jar into the plugins directory of ImageJ.

4. Copy the dependencies into the plugins directory (or onto the Java classpath). Note that the Maven package routine puts all dependencies into the target/dependencies directory even if they are not required by the SMLM code (it does not check what functions are actually used by the code). The libraries required are:

    gdsc-analytics
    gdsc-core
    commons-rng-client-api
    commons-rng-core
    commons-rng-simple
    commons-rng-sampling
    ejml
    JTransforms
    JLargeArrays
    protobuf-java
    protobuf-java-util
    mfl-core
    xstream
    xpp3_min
   

Libraries required if not using the Fiji distribution of ImageJ:

    commons-math3
    commons-lang3
    trove4j
    guava

5. To enable use of the 3D viewer also add the Java3D libraries and the native libraries for your platform. This is unnecessary if using Fiji as the correct libraries are present. The libraries required are:

    3D_Viewer
    j3dcore
    j3dutils
    jogl-all
    jogl-all-*-natives-[platform]
    gluegen-rt
    gluegen-rt-main
    gluegen-rt-*-natives-[platform]
    vecmath-*-scijava-2.jar
    VIB-lib
   

6. The plugins will now appear under the 'Plugins > GDSC SMLM' menu in ImageJ.

Maven can be used to run ImageJ using a profile defined in the gdsc-ij-parent POM:

    mvn -P run-imagej

This profile compiles all classes and then executes ImageJ with the appropriate Java classpath. A default plugin can then be run from the ImageJ Plugins menu to load the plugins defined in the project's ImageJ plugins.config file.

Note: This file is normally detected by ImageJ when loading plugin jar files to identify the available plugins. The default plugin has been written to duplicate this functionality by reading the configuration and populating the ImageJ menu.

The gdsc-smlm code was developed using the Eclipse IDE.

Details of how to open the source code with Eclipse can be found in the eclipse folder.

See LICENSE

Alex Herbert

Genome Damage and Stability Centre, University of Sussex

GDSC SMLM ImageJ plugins