Neutral red

Neutral red is a vital stain used in cell biology and histology for staining lysosomes in living cells. It is a weakly basic dye that is taken up by cells and accumulates in lysosomes, where it is protonated and trapped. Neutral red is commonly used in cytotoxicity assays, such as the Neutral Red Uptake (NRU) assay, to assess cell viability.

Chemical Properties

Neutral red is a member of the phenazine family of dyes. It is a dark green crystalline powder that is soluble in water, forming a red solution. The dye has a molecular formula of C₁₅H₁₇N₄Cl and a molar mass of 288.78 g/mol. Its structure consists of a phenazine core with amino and dimethylamino substituents.

Applications

Neutral red is primarily used in biological research for staining and viability assays. Its applications include:

• Vital Staining: Neutral red is used to stain living cells, particularly to highlight lysosomes. It is a vital stain, meaning it can be used on living cells without killing them.

• Cytotoxicity Assays: The Neutral Red Uptake (NRU) assay is a common method for assessing cell viability and cytotoxicity. In this assay, viable cells incorporate and retain the dye, while dead cells do not.

• Histology: In histological studies, neutral red can be used to stain tissues, providing contrast to highlight cellular structures.

Mechanism of Action

Neutral red is a weak base that penetrates cell membranes by non-ionic diffusion. Once inside the cell, it accumulates in lysosomes due to the acidic environment, where it becomes protonated and trapped. This property makes it useful for identifying lysosomal activity and integrity.

Safety and Handling

Neutral red should be handled with care, as with all chemical reagents. It is important to use appropriate personal protective equipment (PPE) such as gloves and goggles when handling the dye. Neutral red solutions should be prepared in a fume hood to avoid inhalation of dust or vapors.

Also see

• Vital stain
• Lysosome
• Cytotoxicity
• Histology
• Phenazine