Solarbio Weigert’s Gram Stain Solution(Conn Modified)

Specification: 4*50ML

Storage: Room temperature, avoid light

Product Description:

• Gram Staining:
  • Invention:
    • Invented by Danish physician Christian Gram in 1884.
  • Method:
    • Widely used differential staining method in bacteriology.
    • Form of counterstaining.
  • Purpose:
    • Differentiates bacteria into two major classes:
      • Gram-positive (G*)
      • Gram-negative (G)
  • Mechanism:
    • Differential staining based on differences in:
      • Permeability of cell wall to ethanol
      • Ability to resist decolorization
      • Thickness and structure of peptidoglycan layer
  • Staining Process:
    • Cells stained with crystal violet and then treated with iodine form insoluble complexes.
    • Decolorized by ethanol.
  • Isoelectric Points:
    • Gram-positive bacteria: isoelectric points at pH 2.0-3.0
    • Gram-negative bacteria: isoelectric points at pH 4.0-5.0
  • Binding Properties:
    • Gram-positive bacteria carry more negative charges.
    • Bind more tightly to basic dyes such as crystal violet.
  • Formation of Complex:
    • After addition of mordant (iodine) into bacterial cells:
      • Crystal violet-iodine-protein complex insoluble in water is formed.
      • Binds to ribonucleic acid magnesium salts inside Gram-positive bacteria.
  • Result:
    • Stained bacteria less prone to decolorization.

Operating Steps (for reference only):

1. Tissue fixation in 10% formalin, routine dehydration embedding.
2. Cut sections to a thickness of 4μm, and dewax routinely to water.
3. Drop staining slices in crystal violet staining solution for 5 minutes, then discard the dye.
4. Differentiate in acidic ethanol differentiation solution for 2-5 seconds.
5. Rinse with flowing water for 3-5 seconds.
6. Drop staining slices in ammonium oxalate crystal violet staining solution for 5 minutes, then discard the dye. Use paper to gently absorb excess liquid around the slices.
7. Treat slices with Weigert’s iodine solution for 1 minute, then discard the iodine solution. Repeatedly absorb the moisture inside the slices with filter paper.
8. Differentiate with the ortho-toluidine-xylene solution until there is no purple color leaching out of the slices. Immediately wash with xylene, and observe under a microscope. Repeat washing with fresh xylene several times to thoroughly remove orthotoluidine. Seal with neutral resin.

Staining Results:

• Gram-positive bacteria and cellulose: Blue-purple
• Gram-negative bacteria: Red
• Cell nucleus: Red