First immunohistochemical localization of the endogenous Fe2+-chelator nicotianamine

Antibodies produced against nicotianamine-keyhole limpet haemocyanin (NA-KLH) conjugate selectively labelled cells of the stele in tomato root tips (Lycopersicon esculentum Mill. cv. Bonner Beste), where labelling was mostly confined to vacuoles. In competition ELISA this antibody preparation shows no cross-reactivities with precursors for nicotianamine (NA), L-methionine, s-adenosyl-L-methionine, and azetidine-2-carboxylic acid. The antibodies against NA recognize free and metal-bound NA. The usefulness of fixation of NA by glutaraldehyde as a bifunctional reagent is checked by dot blot experiments. The fixation and embedding procedure gave excellent ultrastructural preservation of the cells. The combination of the embedding procedure with the specificity of the used antiserum, the absence of labelling of the NA-free mutant chloronerva, and this lack in immunocytochemical controls, give evidence that it is possible to monitor the NA distribution in situ. Based on this first report on the cellular localization of NA, a low molecular weight iron chelator in plants, the possible roles of NA in mineral metabolism are discussed.