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Electron Microscopy Sciences

Technical Data Sheets

Wolbach's Giemsa Method

EMS Catalog #: 26132-Series

Fixation: 

Russell’s Zenker Fixative (#64123-05) or other well fixed tissues.   

Sections:

Paraffin at 6 µm sections

Staining:

  1. Deparaffinize and hydrate to distilled water.
  2. If Zenker fixed, remove “Zenker’s crystals” by placing in Gram’s Iodine (#26131-04) or Lugol’s Iodine (#26131-4A) for 15 minutes. Rinse in water and clear in Sodium Thiosulfate, 5% (#26131-05) for three minutes. Wash in running water for 15 minutes.
  3. Rinse in distilled water and stain in Working Giemsa Solution (#26132-1A) overnight.

    Working Giemsa Solution may be made from the stock solution (#26132-01) by mixing
    - Giemsa Stock Solution - 1.25 ml
    - Methanol - 1.5 ml
    - Distilled Water - 50.0 ml
    NOTE: Giemsa stain colors more effectively in tissue at an acid pH. If this has not occurred in the preparation or decalcification steps, wash in acid alcohol, then begin stain
  4. Differentiate in Rosin Alcohol Working solution (#26132-2A) until the sections are purplish-pink color. Check under microscope.
  5. Dehydrate in two changes of absolute alcohol and clear in two changes of xylene.
  6. Mount with Permount (#17986-01).

Stain Results:

Nuclei, Bacteria Blue
Rickettsia Purple
Collagen, other tissue elements Pink to rose

References:

Wolbach, S.B., Todd, J.L. and Palfrey, F.W., The Etilogy of Pathology Typhus, Harvard University Press, Cambridge, MA., p.13-14, c. 1922

Luna, L.G., (ed) Manual of Histologic Staining Methods of the AFIP, 3rd edition, McGraw-Hill, New York, p. 119, c. 1968.

Product Information

Wolbach's Giemsa Method