The aluminum-formaldehyde (ALFA) histofluorescence method for improved visualization of catecholamines and indoleamines. 2. Model experiments

The fluorescence-promoting effects of sodium, magnesium and aluminum ions in the histochemical formaldehyde (FA) reaction with catecholamines and indoleamines have been studied in protein models. The positive effects of aluminum and magnesium salts on the monoamine fluorescence yields, seen in tissue, could partly be reproduced in a simple protein matrix. The fluorescence-promoting potency was greatest for aluminum, moderate for magnesium and small for sodium. The aluminum effect was markedly concentration-dependent, with an optimum at a concentration of 5-10 mM in the model solution. Optimum pH was around 3.8. Experiments with synthetic fluorophores indicate that the principal action of the metal ions is on the fluorescence properties of the monoamine fluorophores rather than on the yield of fluorophores in the reaction of the monoamines with FA. The presence of aluminum ions in the models thus causes both increases in the fluorescence efficiency of the fluorophores as well as changes in their spectral properties. The direct effects on the monoamine fluorophores cannot, however, account for all effects of the metal salts seen in tissue. It is suggested that aluminum and magnesium ions, in addition, can act as acid catalysts in the FA-monoamine reactions, and that the metal salts have a direct effect on the tissue environment, leading to a 'locking-in' of the intracellular monoamines.