Fluorescence Microscopy of Aniline Blue Stained Pistils

Abstract

Fluorescence microscopy after aniline blue staining has been used for years to detect “callose-like” substances in plant tissues. Callose-like substances can be polysaccharides or glycoproteins—polysaccharide chains of glucose residues covalently linked to one another by ß1 –> 3 glycosidic bonds are important. Aniline blue is absorbed by such compounds and after exposure to UV light of 430 nm, it emits a yellow–green fluorescence. The method is not chemically specific and many glycoproteins or proteoglycans can be visualized by it (1). Fluorescence is strong and bright only at pH 9.0–10.0. Every change of acidity in the sample has a strong influence on the resulting fluorescence, which can become pale or disappear. The method cannot be taken for quantitative estimation of the callose content, but is sufficient for the morphological or histological investigations applied to anatomy and embryology of plants (2–5).