STAINING

• Staining


  Imparts an artificial coloration to the smear to allow microscopic inspection
• Categories of stains


  • Simple stains
  • Differential stains
  • Probe-mediated stains
• Simple stains


  • Forms and shapes
• Differential stains


  • Specific components of those elements present
• Probe-mediated stains


  • Diagnostic antibody or DNA probe-mediated stains specifically for identification of organisms
• Staining is a technique used to enhance contrast in samples, generally at the microscopic level
• Stains that should be available in all diagnostic microbiology laboratories

  • Gram stain
  • Acid-fast stain
  • Calcofluor white
  • Rapid modified Wright-Giemsa stain
• Gram Stain


  Most commonly used stain in clinical microbiology laboratory
• Gram-positive bacteria


  • Stain dark blue to blue-black with the primary stain crystal violet
• Gram-negative bacteria


  • Stain bright red with the counterstain safranin
• Acid-Fast stain


  Used to stain bacteria that have a high lipid and wax content in their cell walls and do not stain well with traditional bacterial stains
• Ziehl-Neelsen acid-fast stain


  1. Fix smear by heat

  2. Carbolfuchsin, steam gently for 5 min

  3. Wash with deionized water

  4. Decolorize in 3.0% acid-alcohol

  5. Wash with water

  6. Counterstain with Loeffler's methylene blue

  7. Wash with deionized water
• Other common stains used for microscopic evaluation

  • Auramine-rhodamine
  • Acridine Orange
  • Calcofluor White
  • Methylene Blue
  • Lactophenol Cotton Blue
  • India Ink
  • Endospore Stain
  • Rapid Modified Wright-Giemsa Stain
• Calcofluor White


  Fluorochrome that binds to chitin in fungal cell walls, resulting in bright apple-green or blue-white fluorescence
• India Ink


  Negative stain used to visualize capsules surrounding certain yeasts, such as Cryptococcus spp.
• Rapid Modified Wright-Giemsa Stain


  A neutral dye; a combination of basic thiazine dye and acid eosin, resulting in metachromatic staining