Historically, neuroscientists have used microscopes to identify different cell types and determine their distribution in the nervous system. The guidelines, tips, and proposed methodology outlined here, will support researchers of wide-ranging experience levels and areas of focus in neuroscience research. This framework is intended to be simple and yet flexible, such that it can be adapted to suit distinct project needs. To validate our approach, we quantified neuronal density in the mouse barrel cortex using antibodies against pan-neuronal and interneuron markers. Our proposed framework spans from principles and best practices of experimental design, histological and immunofluorescence staining, and microscopy imaging to recommendations for statistical analysis and data presentation. In the present methodological article, we provide a compendium of resources and a FIJI-ImageJ-based workflow aimed at improving the quantification of cell density in mouse brain samples using semi-automated open-science-based methods. Additionally, it is important to consider open science principles (e.g., open-source software and tools, user-friendliness, simplicity, and accessibility). Widespread implementation of many of the elegant image processing tools available continues to be impeded by the lack of complete workflows that span from experimental design, labeling techniques, and analysis workflows, to statistical methods and data presentation.
The ever-expanding availability and evolution of microscopy tools has enabled ground-breaking discoveries in neurobiology, particularly with respect to the analysis of cell-type density and distribution. 8Department of Neurology and Neurosurgery, Centre for Research in Neuroscience, Brain Repair and Integrative Neuroscience Program, Research Institute of the McGill University Health Centre, Montreal General Hospital, Montreal, QC, Canada.7Department of Cellular and Physiological Sciences, University of British Columbia, Vancouver, BC, Canada.6Djavad Mowafaghian Centre for Brain Health, University of British Columbia, Vancouver, BC, Canada.5Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, BC, Canada.4Department of Molecular Medicine, Université de Laval, Québec City, QC, Canada.3Department of Neurology and Neurosurgery, McGill University, Montréal, QC, Canada.2Axe Neurosciences, Centre de Recherche du CHU de Québec, Université de Laval, Québec City, QC, Canada.1Division of Medical Sciences, University of Victoria, Victoria, BC, Canada.Frederiksen 1, Olga Shevtsova 1, Chloe McKee 1, Emma van der Slagt 1, Elisa Gonçalves de Andrade 1, Hai Lam Nguyen 1, Penelope A. Sanchez-Arias 1*†, Micaël Carrier 1,2*†, Simona D.
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