Pituitary and Pancreas 291 



Phloxine: 



phloxine B, C.l. 45410 0.5 gm. 



distilled water 100.0 ml. 



procedure: 



1. Deparaffinize and hydrate slides to water; remove HgCl2. 



2. Refix in Bouin's solution: 12-24 hours. 



3. Wash in running water: 5 minutes. 



4. Treat with potassium dichromate-sulfuric acid: 5 minutes. 



5. Decolorize in sodium bisulfite, 5% (5 gm./lOO ml. water): 3-5 

 minutes. 



6. Wash in rimning water: 5 minutes. 



7. Stain in hematoxylin solution until beta cells are deep blue. 

 Check inider microscope. 10-15 minutes. 



8. Differentiate in hydrochloric acid, 1% (1 ml./99 ml. water): 

 about 1 minute. 



9. Wash in running water luitil clear blue: 5 minutes. 



10. Stain in phloxine: 5 minutes. 



1 1. Rinse briefly in distilled water. 



12. Treat Avith phosphotungstic acid, 5% (5 gm./lOO ml. water): 1 

 minute. 



13. Wash in running water: 5 minutes. Sections turn red again. 



14. Differentiate in 95% alcohol. If the sections are too red and the 

 alpha cells do not stand out clearly, rinse 15-20 seconds in 80% 

 alcohol. 



15. Dehydrate in absolute alcohol, clear, and mount. 



results: 



beta cells — blue 

 alpha cells — red 



delta cells (not present in all animals) — pink to red, actually indis- 

 tinguishable from alphas 



comments: 



Also see Heidenhain's Azan method, page 149. 



If the zymogen granules (acidophilic) are to be preserved in the 

 acinar cells of the pancreatic lobules avoid fixatives containing acetic 

 acid. 



For fluorescent differentiation of the alpha and beta cells, see 

 Hartroft {1951). 



