What are stains ? Types of staining.

What are stains?

• Stains (or dyes) are chemical substances (usually organic salts) used in microbiology to impart color to microorganisms or cell components, making them visible under a microscope.

• They help to highlight structures, differentiate organisms, and study morphology and physiology.

Principle of Staining

• Most stains are salts with a colored ion (chromophore).

• Depending on whether the cation (+) or anion (–) carries the color, stains are classified as:

  • Basic stains: positively charged chromophore (e.g., methylene blue, crystal violet, safranin). These bind to negatively charged cell components (like nucleic acids, cell walls).

  • Acidic stains: negatively charged chromophore (e.g., eosin, nigrosin). Cell surfaces repel these and stain the background (negative staining).

Types of Staining Techniques

A. Simple Staining

• Uses a single dye (e.g., methylene blue, crystal violet).

• Shows the size, shape, and arrangement of microorganisms.

B. Differential Staining

• Uses two or more dyes to differentiate between types of microorganisms or cell structures.

• Examples:

  1. Gram staining differentiates bacteria into two categories: Gram-positive (purple) and Gram-negative (pink/red).

  2. Acid-fast staining (Ziehl–Neelsen stain) – detects Mycobacterium spp. (acid-fast = red; non–acid-fast = blue).

C. Special (Structural) Staining

• Highlights specific structures of microbes.

• Examples:

  1. Capsule stain – negative staining with India ink or nigrosin.

  2. Spore stain (Schaeffer–Fulton method) – spores = green, vegetative cells = red/pink.

  3. Flagella stain – makes flagella visible using special mordants.

D. Negative Staining

• Uses acidic stains (e.g., nigrosin, India ink).

• The background is stained while the cell remains colorless.

• Useful for visualizing capsules and observing bacteria without the need for heat fixation.

In Short:

• Stains = dyes that color microbes or their parts.

• Types of staining =

  1. Simple (one dye, morphology).

  2. Differential (Gram, Acid-fast).

  3. Special/Structural (spores, capsules, flagella).

  4. Negative (background staining).