Overview
This lecture reviews the main types of differential (diagnostic) stains used in microbiology to identify and characterize microorganisms based on their structural features.
Differential Stains Overview
- Differential stains provide information about microbial cell structure, not just color for visibility.
- Common differential stains include Gram, acid-fast, spore (endospore), flagella, and capsule stains.
Gram Stain
- The Gram stain is the first and most important stain used to classify an unknown organism.
- Gram-positive cells have thick peptidoglycan walls and appear purple after staining.
- Gram-negative cells have thin peptidoglycan walls and a lipid outer membrane, appearing red or pink.
- Gram stain method: know the specific stains and their order (details covered in a separate video).
Acid-Fast Stain
- Acid-fast stain identifies organisms with waxy cell walls containing mycolic acid.
- Mainly used for Mycobacterium (e.g., M. tuberculosis, M. leprae) and Nocardia species.
- Acid-fast positive cells stain red; negative cells stain blue.
- Typical stains: Kinyoun’s carbol fuchsin, acid-alcohol decolorizer, and methylene blue counterstain.
Spore (Endospore) Stain
- Used to detect spore-forming bacteria, especially Bacillus and Clostridium genera.
- Spores appear green due to malachite green staining; vegetative cells counterstained with safranin.
- Useful for identifying dangerous spore-formers like C. difficile, C. tetani, and B. anthracis.
Flagella Stain
- Highlights bacterial flagella, which are used for movement.
- A mordant is used to thicken flagella so they become visible under the microscope.
- Helps to distinguish motile organisms.
Capsule Stain (Negative Stain)
- Capsule stains cannot directly color the capsule, so the background and cell are stained instead.
- Capsules appear as clear halos around stained cells due to the negative staining technique.
Key Terms & Definitions
- Differential stain — a stain that distinguishes between cell types or parts based on chemical or structural differences.
- Gram-positive — bacteria with thick peptidoglycan cell walls that stain purple.
- Gram-negative — bacteria with thin peptidoglycan walls and outer membrane, stain red/pink.
- Acid-fast — describes bacteria with waxy cell walls (mycolic acid) that retain certain stains despite acid-alcohol wash.
- Spore (endospore) — a tough, dormant structure formed by some bacteria for survival.
- Mordant — a substance used to fix or intensify stains on cells.
- Negative stain — stains the background, not the target organism or structure.
Action Items / Next Steps
- Study the specific staining procedures for the Gram stain.
- Review related videos on Gram stain and bacterial anatomy.