https://doi.org/10.4081/ejh.2024.3929
Walking a thin line between fixation and epitope binding – characterization of antigen retrieval methods suitable for eosinophil and HSV-2 staining in formalin-fixed female reproductive tissue
Submitted: 29 November 2023
Accepted: 19 March 2024
Published: 16 April 2024
Accepted: 19 March 2024
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All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article or claim that may be made by its manufacturer is not guaranteed or endorsed by the publisher.
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Antibody-based fluorescence analysis of female reproductive tissues in research of sexually transmitted diseases allows for an in-depth understanding of protein localization, interactions, and pathogenesis. However, in many cases, cryosectioning is not compatible with biosafety regulations; at all times, exposure of lab personnel and the public to potentially harmful pathogens from biological infectious material must be avoided; thus, formaldehyde fixation is essential. Due to formaldehyde’s cross-linking properties, protein detection with antibodies can be impeded. To allow effective epitope binding during immunofluorescence of formalin-fixed paraffin-embedded vaginal tissue, we investigated two antigen retrieval methods. We tested these methods regarding their suitability for automated image analysis, facilitating reproducible quantitative microscopic data acquisition in sexually transmitted disease research. Heat-based retrieval at 80°C in citrate buffer proved to increase antibody binding to eosinophil protein and HSV-2 visibly and tissue morphology best, and was the most efficient for sample processing and quantitative analysis.
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Ethics Approval
All protocols and experiments were approved by the Landesamt für Natur, Umwelt und Verbraucherschutz Nordrhein-Westfalen, Recklinghausen, GermanySupporting Agencies
German Research Foundation (DFG) within the German-African Projects in Infectiology, German Center for Infection Research (DZIF), Bundesministerium fur Bildung und ForschungHow to Cite
Wadephul, L. M., Arndts, K., Katawa, G., Dietlmeier, E., Horsnell, W., Hoerauf, A., & Ritter, M. (2024). Walking a thin line between fixation and epitope binding – characterization of antigen retrieval methods suitable for eosinophil and HSV-2 staining in formalin-fixed female reproductive tissue. European Journal of Histochemistry, 68(2). https://doi.org/10.4081/ejh.2024.3929
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