A simple, rapid, and reliable TLC method for the separation and determination of sanguinarine has been established. This intensively studied biologically active alkaloid has a wide range of potentially useful medicinal properties, such as antimicrobial, antiinflammatory, and antitumoral activities. Sanguinarine has also been incorporated into expectorant mixtures and has a strong bactericidal effect upon gram-positive bacteria, particularly Bacillus anthracis and staphylococci. These medicinal properties are due to the interaction of sanguinarine with DNA. A fibre-optic-based fluorescence instrument for in situ scanning was used for quantitative measurements. The sanguinarine was determined over the range 5–40 ng and a detection limit of 1.60 ng. The method was applied to the quantification of sanguinarine in tissue culture extracts of Chelidonium majus L.