366 Special Procedures I (chap. 22) 



Guard (1959) 



fixation: Fix smears immediately in 9% alcohol; do not allow them 

 to dry: 10 minutes. 



solutions: 



Biebrich scarlet: 



biebrich scarlet (water soluble) C.I. 26905 .... I.O gm. 



phosphotungstic acid 0.3 gm. 



glacial acetic acid 5.0 ml. 



50% ethyl alcohol 100.0 ml. 



Fast green: 



fast green FCF C.I. 42053 0.5 gm. 



phosphomolybdic acid 0.3 gm. 



phosphotungstic acid 0.3 gm. 



glacial acetic acid 5.0 ml. 



50% ethyl alcohol 100.0 ml. 



Dilute hematoxylin: 



Harris hematoxylin (page 125) 0.05 ml. 



50% ethyl alcohol 100.0 ml. 



procedure: 



Short. Method: 



1. Transfer into 70% alcohol: 2 minutes. 



2. Stain in biebrich scarlet: 2 minutes. 



3. Rinse in 50% alcohol. 



4. Differentiate in fast green: 1-4 hours. Check under microscope at 

 hourly intervals for green cytoplasm and green vesicular nuclei in 

 all cells. 



5. Rinse in 50% alcohol; 5 minutes. 



6. Dehydrate, clear, and mount. 



Long Method: 



1. Transfer to 70% alcohol: 2 minutes. 



2. Stain in dilute hematoxylin: 15 seconds, or dip in solution 10 

 times. 



3. No rinse, stain in Biebrich scarlet: 2 minutes. 



4. Rinse in 50%, alcohol. 



5. Stain in fast green: 18-24 hours; no microscopic checking neces- 

 sary. 



6. Dehydrate, clear, and mount. 



