Houaloxylin Staining Procedures 137 



procedure: 



1. Deparaffinize and run slides down (hydrate) to water, removing 

 HoCl.. 



2. Oxidize in potassium permanganate, freshly prepared: 5 minutes. 



3. Wash in running water: 5 minutes. 



4. Bleach in oxalic acid: 5 minutes. 



5. Wash in running water: 5 minutes: follow with rinse in distilled 

 ^vater. 



6. Mordant in iron alum: 1 hour. 



7. \Vash out excess alum in running water 10 minutes or longer, 

 rinse in distilled water. 



8. Stain in phosphotungstic acid hematoxylin: 2-24 hours. 

 Usually complete in 2-3 hours, rarely requires overnight. 



9. Transfer directly to 95% alcohol: brief rinse. 



10. Dehydrate, absolute alcohol; watch that elements stained reddish 

 brown are not destained. 



11. Finish dehydration in isopropyl alcohol (99%), 2 changes: 3 

 minutes each. 



12. Clear and mount. 



results: 



nuclei, mitochondria, centrioles, fibrin, fibroglia, myoglia and neu- 

 roglia fibrils, and contractile elements of striated muscle — blue 

 collagen, reticulum, elastin, cartilage and bone— yellowish to brown 

 red 



comments: 



Mullen and McCarter (1941) use a chromium-chloride mordant after 

 formalin fixation: Deparaffinize and hydrate to water, mordant for 2 

 hours or longer in 



chromium chloride, CrCIs 5.0 gm. 



distilled water 100.0 ml. 



glacial acetic acid 5.0 ml. 



Good for several weeks. After this mordanting, the staining will be as 

 brilliant as following Zenker fixation. 



Weigert's Iron Hematoxylin, (mallory 1944; janssen and lillie) 

 fixation: any general fixative or one specific for nuclei. 



solutions: 



Hematoxylin, page 126. 

 Coimterstains, page 129. 



