Title: Wheat Germ Agglutinin (WGA) Staining Optimized for Image Analysis of Muscle Tissue Morphometry 

Authors: Cheru, R. and Wolf, J.C., Experimental Pathology Laboratories, Inc., Sterling, Virginia

Abstract: Wheat Germ Agglutinin (WGA) is a plant-derived lectin and fluorescent stain that binds to N-acetylglucosamine and sialic acid residues in tissues, making it a valuable histochemical tool for visualizing cell membranes and components of the extracellular matrix. In muscle tissue, WGA staining allows clear delineation of the laminin-labeled basal membrane outlining each myofiber, distinguishing it from the residual autofluorescence of the myofiber sarcoplasm. To support digital pathology applications, a WGA staining protocol was optimized for compatibility with image-based quantitative analysis. Formalin-fixed, paraffin-embedded muscle sections were stained with fluorescently labeled WGA, counterstained with DAPI for nuclear visualization, and mounted with antifade medium to preserve fluorescence. Image analysis of WGA-stained skeletal muscle was successfully performed by a pathologist using Image-Pro® Plus software, employing macros to assess myofiber size and count.