Tissue sample Zenker or Helly fixed tissue is recommended, but many fixatives may be used, although staining results may vary. 3µ paraffin sections of neutral buffered formalin fixed tissue are usually suitable.
Method
Bring sections to water with xylene and ethanol.
Rinse well with distilled water.
Place into working Giemsa for 1 hour.
Place into fresh working Giemsa overnight.
Rinse well with distilled water.
Differentiate with working colophonium solution until colour contrast is satisfactory.
Rinse well with 95% ethanol.
Dehydrate with absolute ethanol.
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.
Reference Humason, G.L., (1967) Animal Tissue Techniques
W.H. Freeman & Co., San Francisco, CA, USA
Putt, F.A., (1972) Manual of histopathological staining methods
John Wiley & Sons, New York, NY, USA
