Melzer's is used by exposing fungal tissue or cells to the reagent, typically in a microscope slide preparation, and looking for any of three color reactions:
* Amyloid or Melzer's-positive reaction, in which the material reacts blue to black.
* Pseudoamyloid or dextrinoid reaction, in which the material reacts brown to reddish brown.
* Inamyloid or Melzer's-negative, in which the tissues do not change color, or react faintly yellow-brown.
Among the amyloid reaction, two types can be distinguished:
* Euamyloid reaction, in which the material turns blue without potassium hydroxide (KOH)-pretreatment.
* Hemiamyloid reaction, in which the material turns red in Lugols solution, but shows no reaction in Melzers reagent; when KOH-pretreated it turns blue in both reagents (hemiamyloidity).
Melzer's reactions are typically almost immediate, though in some cases the reaction may take up to 20 minutes to develop.
The function of the chemicals that make up Melzers reagent are several. The chloral hydrate is a clearing agent, bleaching and improving the transparency of various dark-colored microscopic materials. The potassium iodide is used to improve the solubility of the iodine, which is otherwise only semi-soluble in water. Iodine is thought to be the main active staining agent in Melzers; it is thought to react with starch-like polysaccharides in the cell walls of amyloid material, however, its mechanism of action is not entirely understood. It has been observed that hemiamyloid material reacts differently when exposed to Melzers than it does when exposed to other IKI solutions such as Lugols, and that in some cases an amyloid reaction is shown in material that had prior exposure to KOH, but an inamyloid reaction without such pretreatment.
An experiment in which spores from 35 species of basidiomycetes were tested for reactions to both Melzers and Lugols showed that spores in a large percentage of the species tested display very different reactions between the two reagents. These varied from being weakly or non-reactive in Lugols, to giving iodine-positive reactions in Lugols but not in Melzers, to even giving dextrinoid reactions in Lugols while giving amyloid reactions in Melzers.
Melzers degrades into a cloudy precipitate when combined with alkaline solutions, hence it cannot be used in combination or in direct series with such common mycological reagents such as potassium hydroxide or ammonium hydroxide solutions. When potassium hydroxide is used as a pretreatment, the alkalinity must be first neutralized before adding Melzers.