NEUROMOL BIOIMAGE ANALYSIS TOOLS
Microscopy-based data is getting bigger and more complex, which has led to the birth of the research field focused on the development of tools for the analysis of these data. In our lab, we have deployed a series of bioimage analysis workflows with the aim to apply these techniques to the neural stem cell research. Most of the tools are implemented as ImageJ macros, although we also use other open-source software (e.g., CellProfiler, ilastik). All the workflows are freely available to be used and, as open-source software, to be modified at your convenience. Fiji macros can be easily downloaded and added to the plugin menu just by adding the NeuroMol update site, while some CellProfiler pipelines and ilastik projects are also provided via Github (please find the documentation on the corresponding link):
Cell Adhesion Assay
The repository includes a bioimage analysis workflow for the analysis of high throughput microscpy experiments to assess cell adhesion (specifically, to quantify the binding of fluorescence-labelled cells to a cell monolayer) using Fiji.
Cell Proliferation and Apoptosis
The repository includes a bioimage analysis workflow for the analysis of high throughput microscpy experiments to assess cell proliferation in pulse-chase experiments (with BrdU or EdU) using Fiji. Additionally, the macro allows to quantify the signal of up to 2 extra nuclear markers, so the workflow can be applied either to label different cell subpopulations or to look for additional information in order to assess the cell culture viability (e.g., counting apoptotic events by labelling caspase3-positive cells).
Neurosphere Formation Assay
The repository includes a bioimage analysis workflow for the assessment of the clonal capacity and self-renewal on spheroids (e.g., neurospheres). It consists on a set of Fiji macros to be run sequentially and connected to ilastik.
The repository contains a series of CellProfiler pipelines aimed to quantify the cyt-nuc ratio of a given marker on microscopy-based screening assays. These pipelines were deployed during a Short Term Scientific Mission founded by NEUBIAS and hosted by the Advanced Light Microscopy (ALM) platform and the BioScreening platform at the Instituto de Investigação e Inovação em Saúde (i3S, Universidade do Porto).
This is an ongoing project which aims to accomplish the segmentation of three axonal components (axoplasm, inner cytoplasmic tongue and myelin) on TEM images using Fiji and ilastik. The project is a collaboration with the Anna Williams’s Lab (MRC Centre for Regenerative Medicine, University of Edinburgh).