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REALM

Open-source Adaptive Optics plugin for Micro-Manager

Introduction

REALM stands for Robust and Effective Adaptive optics in Localization Microscopy and can perform aberration correction in Single Molecule Localization Microscopy (SMLM). It can however also be used for aberration correction on bead(s) and other small fiducial markers or to flatten the deformable mirror on a bead sample.

REALM provides an user-friendly interface to allow users to perform SMLM or other types of microscopy in tissue or other complex samples. It is designed for users with little background adaptive optics, but provides access to important parameters such that advanced users can tune the aberration correction algorithm to their needs.

Deformable mirror support

Currently REALM supports two deformable mirrors: the MIRAO52E from Imagine Optic and the DMP40 from Thorlabs. Micro-Manager uses device adapters to communicate with devices. You can download the device adapter for MIRAO52E from https://github.com/MSiemons/MIRAO_DeviceAdapter. Here a manual is provided on how to install this device adapter. The device adapter for Thorlabs can be found at https://github.com/HohlbeinLab/Thorlabs_DM_Device_Adapter.

Do you have a different DM? Contact me to help you interface with REALM!

Installation

REALM is installed in 4 easy steps.

  1. Download the .zip file from the repository https://github.com/MSiemons/REALM.
  2. Create a folder named “REALM” in the Micro-Manager installation folder, extract the zip-file and copy all files to this folder. Alt text
  3. Copy the latest version REALM-X.jar from the dist-folder to the mmplugins folder.
  4. Start Micro-Manager and check that Micro-Manager recognizes the plugin. REALM can be found under ‘Plugins’. REALM is now installed.

The REALM GUI

The GUI of REALM features a nice big START button and provides acces to the aberration correction parameters in the settings tab.

Correction algorithm

REALM uses model-based optimization to iteratively correct Zernike modes. In brief, each Zernike mode is corrected by applying a sequence of different amounts of biases of the to be corrected Zernike mode. From these acquisitions a metric is computed. Next a Gaussian is fitted through these metric values to find the optimum. This is then performed for all the to be corrected Zernike modes and is repeated for multiple correction rounds.

Modifying REALM

The specific implementation of REALM might not be perfectly suited for your needs. For instance, you prefer a different correction order of the Zernike modes. In REALMframe.java you can edit the specific Zernike mode lists and compile REALM yourself. Please look at https://micro-manager.org/writing_plugins_for_Micro-Manager-1.4 for instructions how to set up the compile environment.

Citing

If you use REALM please cite our paper

Siemons, M.E., Hanemaaijer, N.A.K., Kole, M.H.P. Kapitein, L.C., Robust adaptive optics for localization microscopy deep in complex tissue. Nat Commun 12, 3407 (2021). https://doi.org/10.1038/s41467-021-23647-2