STAINING, PRACTICAL AND THEORETICAL 



3. Rinse for a few minutes in 50% alcohol; then in distilled 

 water. 



4. Counterstain in carmalum for about three minutes; wash 

 with distilled water; mount in glycerine jelly. 



Results: 



Neutral fat and myelin : blue-black to black ; nuclei : red. 



Reference: Carleton, H. M. & Leach, E. H. (1947), p. 259. 



SUDAN BLACK - ETHYLENE GLYCOL 



An improved technique for lipid staining, offering the ad- 

 vantage of a stable solution, excellent differentiation with- 

 out loss of stain out of the lipid particles, and pliable 



unshrunken sections 



Solutions required: 



A. Ethylene glycol, pure, anhydrous. 



B. Sudan black . . . . . . . . i gm. 



Ethylene glycol, pure, anhydrous. . 100 ml. 



Heat the ethylene glycol to 100-110° C. on a hot plate or in an 

 oven, or over a bunsen flame, taking care that it does not catch 

 fire ; then add the stain and stir until all or most of it is dissolved. 

 Filter when cold. 



C. Ethylene glycol, pure, anhydrous . . 85 ml. 

 Distilled water . . . . . . 15 ml. 



D. Carmalum (Mayer). 



Technique: 



1. Fix tissues for at least three days in 10% formalin. 



2. Wash thoroughly in running water; cut frozen sections. 



3. Dehydrate the sections by agitating gently in pure anhydrous 

 ethylene glycol for three to five minutes. 



4. Immerse the sections in the Sudan black solution from 

 fifteen minutes to one hour, agitating gently at intervals. 



5. Differentiate by agitating gently at intervals with 85% 

 ethylene glycol (solution C) from one to ten minutes, controlling 

 under the microscope while the section is still wet. 



428 



