Work

Histological and Immunolabeling Techniques in Arabidopsis thaliana: A Practical Guide and Standardization Roadmap

Year: 2025

Type: article

Abstract: Arabidopsis thaliana is a widely used model in plant biology, where histology (HT), histochemistry (HC), immunohistochemistry (IHC), and immunofluorescence (IF) are applied to study cellular structures, macromolecules, and antigens. Despite their extensive use, protocols lack standardization and exhibit substantial variability in critical aspects such as reagent handling, exposure times, and the proper use of controls. This methodological heterogeneity represents a major gap, limiting reproducibility and comparability between studies. Unlike previous methodological reviews, this work focuses exclusively on A. thaliana, systematically identifies reporting omissions, and proposes a roadmap for standardization. A narrative review of literature retrieved from Scopus and Web of Science was conducted with the aim of analyzing methodological approaches, identifying inconsistencies, and offering recommendations for improved laboratory practices. The analysis revealed frequent omissions in the reporting of critical steps such as dehydration, clearing, antigen retrieval, enzyme blocking, and the incorporation of positive and negative controls, which compromise the reliability of results and hinder inter-laboratory validation. Based on this evidence, three key recommendations are emphasized: (i) organ-specific selection and explicit justification of fixatives and stains; (ii) mandatory incorporation of positive and negative controls in IHC and IF; and (iii) adoption of a minimum reporting checklist to enhance reproducibility. Beyond cell morphology, the reviewed studies demonstrate applications in plant physiology, phytogenetics, and pathophysiology. By combining critical analysis with actionable guidelines, this review contributes a practical reference to strengthen methodological rigor in histological and immunological studies of plants.

Source: Agronomy

DOI: 10.3390/agronomy15102357