SECTION TWO 



8. Wash with water. 



9. Immerse in 50% alcohol for two minutes. 



10. Immerse in 95% alcohol for two minutes. 



11. Immerse in the fast green-orange 2 (Solution C) for five 

 minutes. 



12. Differentiate, examining under the microscope, until the 

 connective tissue is stained to the desired depth of green. 



13. Immerse in clove oil for ten minutes. 



14. Transfer to Orange 2 in clove oil for ten minutes. 



15. Differentiate, examining under the microscope at intervals, 

 until the desired depth of staining has been achieved. 



16. Immerse for ten minutes each in two changes of xylol. 

 12. Mount in balsam or D.P.X. 



Results: 



Nuclei: red. Nucleoli: purple or purplish red. Nuclear mem- 

 brane : dark red. Cellular cytoplasm : pink to red, except in Henle's 

 loop (light green). Connective tissue : green. Elastic fibres : yellow. 

 Fibroblasts: green with purple nuclei. Muscle: reddish brown. 

 Erythrocytes: orange. Polymorphonuclears show purple nuclei. 



Notes: 



The original author (Kalter) states that this technique, which 

 is a development of Flemming's triple stain (Flemming, 1891), is 

 particularly useful for histology students. 



Reference: Kalter, S. S. (1943). 



SAFRANIN - WATER BLUE 



(After Unna) 

 For collagen fibres 



Solutions required: 



A. Safranin O 1% aqueous. 



B. Water Blue 1% aqueous. . . . 10 ml. 

 Tannic acid 33% aqueous . . 10 ml. 



This solution must be freshly prepared. 



397 



