Sechenov School
Home › Histology › Histological Stains: Types and Mechanisms

Histological Stains

For medical students2 min readUpdated 2026-10-10

Histological stains are specialized chemical agents used to visualize microscopic structures of cells and tissues. Depending on their chemical nature and specific affinity for various tissue components, they are divided into four major groups.

GroupsThere are 4 groups of stains based on chemical nature and affinity
AcidicBind to proteins and collagen (the phenomenon of oxyphilia)
BasicStain DNA, RNA, and hyaluronic acid (the phenomenon of basophilia)
IndifferentDissolve in lipids, staining them bright orange

Principles of Classification

To study tissues under a microscope, they must be contrasted. All staining agents used in histology are divided into exactly four groups. The primary criterion for this division is the chemical nature of the stain itself, as well as its affinity for specific structures inside the cell or within the extracellular matrix. Understanding this classification is key to interpreting any histological specimen.

Acidic Stains and Oxyphilia

Chemically, these substances are acids or their acidic salts. The key property of tissue structures that allows them to bind to such substances is called oxyphilia (or acidophilia).

Typical representatives of this group include bright pink eosin and acid fuchsin. Their main "targets" are oxyphilic structures rich in basic chemical groups:

Basic Stains and Basophilia

This group is represented by basic salts. The ability of tissues to bind to them is termed basophilia. Among the most well-known representatives are carmine, azure II, and hematoxylin (with the active coloring agent in the latter being its oxidation product — hematein).

Basic stains selectively bind to structures rich in organic acids. Such basophilic targets include:

  1. Cell nuclei and ribosomes. Staining occurs due to their high content of nucleic acids (DNA and RNA).
  2. The amorphous component of the extracellular matrix. Here, the reaction is due to the presence of hyaluronic acid and similar compounds.

Neutral and Indifferent Stains

Neutral stains represent a complex mixture of basic and acidic coloring agents. Their mechanism of action involves the simultaneous staining of different tissue components in their respective specific colors. Classic targets for them (neutrophilic structures) are the specific granulation of neutrophil granulocytes and the cytoplasm of polychromatophilic erythroblasts.

Indifferent stains differ cardinally from the rest. By nature, they are hydrophobic and lipophilic substances that are completely insoluble in water. The most famous representatives are Sudan III and Sudan IV.

Their staining mechanism is based not on a chemical reaction with tissue components, but on the purely physical dissolution of the stain itself in fats and lipids. As a result, lipid droplets acquire a rich bright orange color, making them easy to identify on a slide.

Mnemonic

Remember basophilia easily: BAsic stains bind to acids (DNA, RNA in nuclei). Oxyphilia is the opposite: ACIdic stains color proteins and cytoplasm.

Frequently asked questions

How do metachromatic stains differ from orthochromatic stains?

Metachromatic stains have the ability to change their original color upon contact with chromotropic substances. Metachromasia is due to a high concentration of glycosaminoglycans in the tissue. Examples of metachromatic stains include:

  • Toluidine blue — stains the substance not blue, but a different color (purple-red).
  • Methylene blue — can also exhibit metachromasia.
Which histological stains are used to detect glycogen?

Glycogen is detected using Schiff's reagent (in the PAS reaction) and Best's carmine.

  • Schiff's reagent — used in the PAS reaction after periodic acid oxidation. Glycogen clumps stain purple, dark red, or dark magenta.
  • Best's carmine — used for specific differentiation of glycogen, giving a red color.

Enzymatic control (amylase digestion of sections) is used to confirm the presence of glycogen.

What stains are used to identify elastic fibers?

Elastic fibers are identified usingorcein or a combination of picrofuchsin and hematoxylin, as they lack contrast with standard methods. Staining methods:

  • Orcein — a selective method where elastic fibers and membranes acquire a cherry (or cherry-red) color against a pale pink background of other tissues.
  • Picrofuchsin with hematoxylin — a differential method that stains elastic fibers yellow due to picric acid, collagen fibers red, and cell nuclei purple.
What is the difference between oxyphilia and basophilia?

Oxyphilia is the ability of structures to stain with acidic stains (such as cytoplasmic proteins and collagen). Basophilia is the affinity for basic stains, characteristic of structures rich in acids (nuclei, ribosomes, amorphous ground substance).

What is the mechanism of action of indifferent stains?

Unlike other groups, they do not enter into chemical reactions with tissue. Indifferent stains (e.g., Sudan III) physically dissolve in lipids and fats, coloring them bright orange.

What do neutral stains color?

They consist of a mixture of acidic and basic substances, staining neutrophilic structures: the specific granules of neutrophil granulocytes and the cytoplasm of polychromatophilic erythroblasts.

Go deeper

More topics in Histology

OvumGastrulation in AmphibiansChorion and AmnionGamete TransportSimple EpitheliaCell Membranes: Structure and FunctionsBlood Groups and Transfusion CompatibilityDense Fibrous Connective TissueSkeletal Muscle TissueNeuronsProprioceptive Sensitivity and Muscle SpindlesSympathetic and Parasympathetic Reflex ArcsHistology →