Amyloid is an amorphous extracellular eosinophilic substance that can be present in various tissues and organs, and the diseases caused by it are referred to as amyloidosis. Amyloid is mainly composed of proteins, most of which are arranged in an antiparallel β-pleated sheet structure. Under an electron microscope, amyloid exhibits a fibrillary arrangement, appearing as a large number of unbranched extracellular filaments in pathological specimens, mostly arranged randomly.
Histological methods for identifying amyloid include methyl violet staining, Congo red staining, and polarized light microscopy. Current research has demonstrated that the traditional methyl violet staining method has low sensitivity and poor specificity, while Congo red staining is a classic and effective approach. In 1922, Bennhold discovered that Congo red could be used for the identification of amyloid in vivo and applied this technique to tissue sections. Later, the method was modified by Highman, resulting in improved staining efficacy.
Amyloid Staining Solution (Methyl Violet Method) is mainly composed of Methyl Violet Staining Solution and Acid Differentiating Solution. It is a metachromatic staining solution modified by Jurgens. Its staining principle is that the acid mucopolysaccharides in amyloid react metachromatically with methyl violet. The advantages of this method are simplicity and time-saving, while its disadvantage is that the stained sections are difficult to preserve. This reagent is for research use only and is not suitable for clinical diagnosis or any other purposes.
A1508465
| Component | 2×50 mL | Storage | | A1508465A | Methyl Violet Staining Solution
| 50 mL | RT. Store in the dark.
| | A1508465B | Acid Differentiating Solution
| 50 mL | RT |
|
Materials to be Prepared by User
10% neutral formalin, distilled water, graded ethanol, xylene or environment-friendly paraffin removal and clearing solution, glycerin gelatin
Procedure (for reference only)
1. Perform routine fixation, usually with 10% neutral formalin fixative, followed by routine dehydration and embedding.
2. Cut sections with a thickness of 4 μm, and perform routine dewaxing to water using xylene or dewaxing and clearing solution.
3. Immerse the sections in Methyl Violet Staining Solution for 3 min without rinsing with water.
4. Add Acid Differentiating Solution dropwise for differentiation until no more stain elutes out.
5. Rinse the sections slightly with water and mount them with glycerin gelatin.
Staining Results
Amyloid: Red to purplish red
Cell nuclei, cytoplasm, connective tissue: Blue to purple-blue of varying shades
Precautions
1. Ensure thorough dewaxing of sections; otherwise, the staining effect will be affected.
2. Prefer immersion staining; if drop staining is adopted, place the sections in a moist chamber to prevent solution evaporation.
3. Store Acid Differentiating Solution in a sealed container. The differentiation step is critical for staining quality.
4. Do not dehydrate the stained sections with ethanol, otherwise, the problem of failure to stain may occur.
5. Mucous substances also show metachromatic red color when stained with methyl violet, so attention should be paid to differentiation.
6. When observing the metachromatic reaction under the microscope, the blue filter should be removed.
7. For your safety and health, wear a lab coat and disposable gloves during operation.
8. Use the reagent as soon as possible after opening to avoid affecting the results of subsequent experiments.
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