SECTION TWO 



C. Giemsa stain . . . . . . 0-25 ml. 



Distilled water . . . . . . 100 ml. 



Note: Solution C should be prepared freshly, as 

 required, from Giemsa stain. 



D. Alcohol 95% 100 ml. 



Eosin 1% in 90% alcohol . . i ml. 



Technique: 



1. Paraffin sections are brought down to 70% alcohol. 



2. Stain for one half to one hour in Solution A; then rinse for 

 two to five minutes in distilled water. 



3. Wipe off excess water; dip into 95% alcohol for a few sec- 

 onds; then decolorize with absolute alcohol for five to twenty- 

 five minutes, or until the sections assume a pale brown colour and 

 the elastic fibres stand out, deep purple to black, under the low- 

 power objective. 



4. Decolorize in Solution B until the background is almost 

 colourless. This usually takes two to seven minutes. 



Note: Decolorization must not be extended more than ten 

 minutes, as otherwise the thin elastic fibres will become destained. 



5. Immerse in tap water for five to ten minutes. 



6. Stain for two to twelve hours with Solution C until the 

 epithelial and other cells are deep blue; connective tissue, greyish 

 pink or greyish blue or blue. 



7. Wipe off excess stain and dehydrate and decolorize in 

 Solution D, controlling under the microscope. 



Note: Decolorization must be stopped when the connective 

 tissue has lost all trace of blue and has assumed a rose tint. The 

 blue tinge is removed fairly rapidly in Solution D. The epidermis 

 should remain bright blue. 



8. Immerse in two changes of absolute alcohol for two minutes 

 in each. Clear in xylol, and mount. 



Results: 



Nuclei: deep blue. Cytoplasm of the epidermis, muscle cells 

 and connective tissue cells : light blue. Plasma cells : dark greyish 

 blue. Eosinophilic granules : bright red. Mast cell granules : meta- 

 chromatic (varying shades of) purple. Neutrophilic granules: only 



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