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A freely available semi-automated method for quantifying retinal ganglion cells in entire retinal flatmounts

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dc.contributor.authorGeeraerts, E.
dc.contributor.authorDekeyster, E.
dc.contributor.authorGaublomme, D.
dc.contributor.authorDe Groef, L.
dc.contributor.authorMoons, L.
dc.contributor.authorSalinas Navarro, Manuel Ángel
dc.contributor.departmentAnatomía Humana y Psicobiología
dc.date.accessioned2026-03-05T09:08:14Z
dc.date.available2026-03-05T09:08:14Z
dc.date.copyright© 2016 Elsevier Ltd.
dc.date.issued2016-04-20
dc.description.abstractGlaucomatous optic neuropathies are characterized by progressive loss of retinal ganglion cells (RGCs), the neurons that connect the eye to the brain. Quantification of these RGCs is a cornerstone in experimental optic neuropathy research and commonly performed via manually quantifying parts of the retina. However, this is a time-consuming process subject to inter- and intra-observer variability. Here we present a freely available ImageJ script to semi-automatically quantify RGCs in entire retinal flatmounts after immunostaining for the RGC-specific transcription factor Brn3a. The blob-like signal of Brn3a-immunopositive RGCs is enhanced via eigenvalues of the Hessian matrix and the resulting local maxima are counted as RGCs. After the user has outlined the retinal flatmount area, the total RGC number and retinal area are reported and an isodensity map, showing the RGC density distribution across the retina, is created. The semi-automated quantification shows a very strong correlation (Pearson's r ≥ 0.99) with manual counts for both widefield and confocal images, thereby validating the data generated via the developed script. Moreover, application of this method in established glaucomatous optic neuropathy models such as N-methyl-D-aspartate-induced excitotoxicity, optic nerve crush and laser-induced ocular hypertension revealed RGC loss conform with literature. Compared to manual counting, the described automated quantification method is faster and shows user-independent consistency. Furthermore, as the script detects the RGC number in entire retinal flatmounts, the method allows detection of regional differences in RGC density. As such, it can help advance research investigating the degenerative mechanisms of glaucomatous optic neuropathies and the effectiveness of new neuroprotective treatments. Because the script is flexible and easy to optimize due to a low number of critical parameters, it can potentially be applied in combination with other tissues or alternative labeling protocols.
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dc.identifier.citationGeeraerts, E., Dekeyster, E., Gaublomme, D., Salinas-Navarro, M., De Groef, L., & Moons, L. (2016). A freely available semi-automated method for quantifying retinal ganglion cells in entire retinal flatmounts. Experimental eye research, 147, 105-113.
dc.identifier.doihttps://doi.org/10.1016/j.exer.2016.04.010
dc.identifier.issn0014-4835
dc.identifier.urihttp://hdl.handle.net/10201/218301
dc.languageeng
dc.publisherElsevier
dc.relationThis work was supported by the KU Leuven Research Council (KU Leuven, Belgium, BOF-OT/14/064), the Hercules Foundation (Belgium, AKUL-09-038, AKUL1309), the Research Foundation Flanders (FWO-Vlaanderen, Belgium, G0B2315N; fellowships to ED, DG and MSN), and the Flemish government agency for Innovation by Science and Technology (IWT-Vlaanderen, Belgium, fellowships to LDG and EG).
dc.relation.publisherversionhttps://www.sciencedirect.com/science/article/pii/S0014483516300781
dc.rights.accessRightsinfo:eu-repo/semantics/restrictedAccess
dc.subjectGlaucomatous optic neuropathies
dc.subjectRetinal ganglion cell
dc.subjectSemi automated quantification
dc.subjectBrn3a
dc.subjectMouse
dc.subject.odsNo relacionado con ningún objetivo de desarrollo sostenible
dc.titleA freely available semi-automated method for quantifying retinal ganglion cells in entire retinal flatmounts
dc.typeinfo:eu-repo/semantics/article
dc.type.versioninfo:eu-repo/semantics/publishedVersion
dspace.entity.typePublicationes
relation.isAuthorOfPublication81da925b-fa46-4a1a-96fd-ce35568fa423
relation.isAuthorOfPublication.latestForDiscovery81da925b-fa46-4a1a-96fd-ce35568fa423
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