Skip to main content

Wolbach’s Giemsa General Oversight Stain

Wolbach's Giemsa

General Oversight Stain

9
steps
7
materials

Materials

  • Stock Giemsa
  • Working Giemsa
    MaterialAmount
    Stock Giemsa2.5mL
    Methanol3mL
    Distilled water100mL

    Make the working solution just before using.

  • Stock Colophonium
    MaterialAmount
    Colophonium10g
    Absolute ethanol100mL

Tissue Sample

Zenker or Helly fixed tissue is recommended, but many fixatives may be used, although staining results may vary. 3 µ paraffin sections of neutral buffered formalin fixed tissue are usually suitable.

Protocol

  1. Bring sections to water with xylene and ethanol.
  2. Rinse well with distilled water.
  3. Place into working Giemsa for 1 hour.
  4. Place into fresh working Giemsa overnight.
  5. Rinse well with distilled water.
  6. Differentiate with working colophonium solution until color contrast is satisfactory.
  7. Rinse well with 95% ethanol.
  8. Dehydrate with absolute ethanol.
  9. Clear with xylene and mount with a synthetic resinous medium.

Expected Results

  • Nuclei – blue-purple
  • Cytoplasm – pink
  • Inclusions – according to type – pink or blue

Notes

  • Humason says to treat sections with 1% acid alcohol for 5 minutes immediately after bringing to water. They are then thoroughly washed with running tap water for 5 minutes. This would have the effect of improving staining by the eosin component of the Giemsa.
  • Putt says to add 3-4 drops of 0.5% aqueous sodium bicarbonate to the water used to dilute the Giemsa. This would have the effect of improving staining by the basic dye component of the Giemsa.
  • Humason notes that the diluted Giemsa may be heated to 80-85°C then poured on the slide and applied for 10 minutes.

Safety Note

Prior to handling any chemical, consult the Safety Data Sheet (SDS) for proper handling and safety precautions.

References

  1. Humason, G.L., (1967)
    Animal Tissue Techniques
    W.H. Freeman & Co., San Francisco, CA, USA
  2. Putt, F.A., (1972)
    Manual of histopathological staining methods
    John Wiley & Sons, New York, NY, USA