Melzer's Reagent, abbreviated as Melzer's, was first invented and described in 1924 by the mycologist Václav Melzer, and named after its inventor. Melzer modified a chloral hydrate-containing KI solution developed many years earlier by the botanist Arthur Meyer to obtain Melzer's Reagent. Melzer's Reagent is often used together with PVLG solution.
Melzer's Reagent consists of chloral hydrate, potassium iodide, iodine, and deionized water. It is of great significance in the amyloidity analysis of spore ornamentation or entire spores, and is used by mycologists to assist in fungal identification. When using Melzer's Reagent, the reagent only needs to contact fungal tissues or cells, and then observe whether one of the following three color reactions occurs:
① Amyloid or Melzer-positive reaction: The tested tissue appears blue to black.
② Pseudoamyloid or dextrinoid reaction: The tested tissue appears brown to reddish-brown.
③ Inamyloid or Melzer-negative reaction: The tested tissue shows no color change, or exhibits pale yellowish-brown.
Two types can be further distinguished in amyloid reactions:
A. Euamyloid reaction: The tested tissue turns blue without pre-treatment with potassium hydroxide.
B. Hemiamyloid reaction: The tested tissue turns red in Lugol's solution but shows no reaction in Melzer's Reagent; after pre-treatment with potassium hydroxide, it turns blue in both reagents.
The reaction of Melzer's Reagent occurs almost instantaneously, although in some cases up to 20 minutes may be required for the reaction to be observed. The compounds composing Melzer's Reagent serve multiple functions. Chloral hydrate acts as a clearing agent, used for bleaching and increasing the transparency of many dark-colored substances. Potassium iodide is used to improve the solubility of iodine; without potassium iodide, iodine is only partially soluble in water. Iodine is considered the primary staining agent in Melzer's Reagent, which reacts with starch-like polysaccharides in the cell walls of amyloid substances.
Materials Required (User-supplied):
Test sample, PVLG solution, potassium hydroxide solution, glass slides, coverslips, microscope, dissecting needle or pipette, forceps
Procedures (for reference only):
Perform operations according to specific experimental requirements.
Staining Results:
Amyloid or Melzer-positive reaction: The tested tissue appears blue to black.
Pseudoamyloid or dextrinoid reaction: The tested tissue appears brown to reddish-brown.
Inamyloid or Melzer-negative reaction: The tested tissue shows no color change, or exhibits pale yellowish-brown.
Euamyloid reaction: The tested tissue turns blue without pre-treatment with potassium hydroxide.
Hemiamyloid reaction: The tested tissue turns red in Lugol's solution but shows no reaction in Melzer's Reagent; after pre-treatment with potassium hydroxide, it turns blue in both reagents.
Precautions:
1. For your safety and health, please wear a laboratory coat and disposable gloves during operation.
2. Most spores exhibit different reaction results in Melzer's Reagent and Lugol's reagent. Some spores show no reaction in Melzer's Reagent but range from weak or no reaction to iodine-positive in Lugol's reagent; some spores show an amyloid reaction in Melzer's Reagent and even a pseudoamyloid reaction in Lugol's reagent.
3. Melzer's Reagent degrades into a turbid precipitate when mixed with alkaline solutions. Therefore, it must not be mixed or used sequentially with common mycological reagents such as potassium hydroxide or ammonia solution. When potassium hydroxide is used for pre-treatment, the alkalinity must be neutralized before adding Melzer's Reagent.
4. When Melzer's Reagent is mixed with PVLG solution at a ratio of 1:1, the staining reaction decreases slightly, but not sufficiently to affect the overall color change of the reaction. Preparations cannot be preserved for a long time, and the staining reaction usually fades within 1–2 years.
5. The purpose of allowing to stand for 5 min before mounting is to slightly dry the reagent, increase edge adhesion, and reduce liquid flow that would cause spores to slide outside the coverslip during mounting.
6. Under the microscope, press directly on the coverslip with the blunt end of a pencil, forceps, or dissecting needle to apply indirect pressure to individual spores. As each spore is compressed to a different degree, the delicate elastic inner walls that are otherwise difficult to observe will be revealed.
7. Please use the reagent as soon as possible after opening to avoid affecting subsequent experimental results.
8. This product is for research use only; strictly prohibited for other purposes.
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