Toluidine Blue O is widely used in biological and medical fields as a vital stain for microscopy. It is particularly effective in staining nucleic acids, making it a valuable tool for identifying cell nuclei in tissue samples. The dye is also employed in histopathology to differentiate between various types of tissues and cells, aiding in the diagnosis of diseases such as cancer. In addition, it is used in microbiology to stain and identify bacteria and other microorganisms. Toluidine Blue O is also utilized in dermatology for detecting malignant lesions, as it selectively stains abnormal tissues. Its metachromatic properties make it useful in studying cartilage and other connective tissues. Furthermore, it is applied in research to investigate cellular structures and functions, providing insights into cellular processes and mechanisms.