Protocol to analyse the structural composition by fluorescence microscopy and different conventional and fluorescence staining methods

The protocol shows the effectiveness of using safranin-fast green stain for fluorescence microscopy. This staining technique has been used in conventional microscopy to perform anatomical characterizations of plants. However, this protocol describes the procedure for using samples stained with safranin-fast green in conjunction with fluorescence microscopy. The strength of the protocol lies in the fact that the samples are permanent and allows for effective differentiation of lignified and cellulosic walls unlike conventional fluorescence microscopy stains such as Congo red-acridine orange, calcofluor, and autofluorescence. The protocol for making fluorescence intensity measurements is also standardized, allowing the data to be used for statistical analysis and inference about the chemical composition of plant cell walls.