Absolute Quantification and Spatial Mapping of Hyaluronic Acid in Histological Tissue Sections

We present an accessible methodology for the absolute quantification and spatial mapping of hyaluronic acid (HA) in paraffin-embedded tissues. Although HA plays critical roles in tissue hydration, organization, and disease progression, its local concentration and spatial distribution remain poorly defined due to the lack of quantitative measurement tools. By integrating immunofluorescence, ELISA, and image analysis our approach enables the generation of pixel-level HA concentration maps. This strategy bridges the gap between qualitative imaging modalities and bulk biochemical assays, offering both spatial resolution and quantitative data. The method is validated across multiple tissues and is robust, scalable, and readily applicable to other extracellular matrix components, offering a practical tool for studying tissue microenvironments in health, disease, and biomaterial design.