96 The Preparation of Microscope Slides 



Stain Is Used as Follows: 



1. Accumulate sections mounted on slides in 70% alcohol and place 

 them in the first staining solution for 1 to 3 days. 



2. Wash slides in running tap water until no more color comes away. 



3. Transfer them to the second staining solution for 10 to 15 min. 



4. Rinse slides in running tap water. 



5. Differentiate them for 10 to 15 sec in the first differentiating solu- 

 tion. 



6. Stain sections in the third staining solution from 10 to 20 min or 

 until sufficient green dye has been absorbed. 



7. Rinse them for 5 to 10 sec in the second differentiating solution. 



8. Place sections in the fourth staining solution from 3 to 5 min or until 

 the cytoplasm of the cells has become bright orange. 



9. Dip them up and down three or four times in the third differentiat- 

 ing solution. 



10. Rinse for 10 to 15 sec in special dehydrating solution. 



11. Transfer slides to xylene and mount in the ordinary manner. 



In a successful preparation of this type, chromosomes and lignified cell 

 walls are stained bright red, the contents of the cells having purplish 

 resting nuclei against a bright-orange cytoplasm. There is a very vivid, 

 but somewhat variable differentiation of the remaining structures. Parasitic 

 fungi are particularly well shown in bright green whether they are pene- 

 trating cytoplasm or the lignified portion of the cell wall. 



STAINS FOR SPECIAL PURPOSES 



The number of stains for special purposes is legion. They belong prop- 

 erly in the field of the specialist in the particular tissue that they demon- 

 strate. There are, however, a few that are sufficiently interesting and 

 simple to justify their inclusion in a beginner's handbook. 



Fats. Most fats are normally dissolved from tissues in the course of 

 embedding in wax. If, however, frozen sections are made by the tech- 

 nique described on page 163, it is possible to demonstrate differentially 

 the fat globules by soaking the sections in an alcoholic solution of a fat- 

 soluble, but water-insoluble, dye. The classic method is to use a saturated 

 solution of sudan IV in 70 per cent alcohol. This stains fat globules red. 

 A blue color may be obtained by using a saturated solution of oil blue N 

 in 60 per cent isopropyl alcohol. Sections stained by this method cannot 

 be dehydrated and should be mounted in Farrants's medium ( see p. 104 ) . 



Skeletons. It is often useful, in the study of embryos or very small 

 vertebrates, to be able to stain the skeleton differentially. 



Specimens, such as fish fry, which have bony skeletons, should be 



