Method XV 89 



11. To stain, replace the water with about 20 volumes of the 

 haematoxylin solution (E), cover, and set aside for 24 hours. 



12. To differentiate and fix the stain, rinse quickly in distilled 

 water, one section at a time. With a needle hold the section by 

 celloidin margin on a section lifter, and merely dip it through 

 the water three or four times, and, still holding the section, 

 transfer it to a dish containing the ferric chloride solution (F). 

 Here the Sections should remain anywhere from 5-20 minutes. 

 To determine the optimum time, expose different sections for 

 varying lengths of time. 



13. From the ferric chloride, wash in distilled water for 5-10 

 minutes. 



14^ To clear and mount, dehydrate quickly by transferring 

 directly to a copious amount of 95 per cent, alcohol, agitating the 

 alcohol about the sections for 20-30 seconds. 



Then to clear, transfer to a covered dish containing a small 

 amount of origanum oil (G). The clearing should take place in 

 5-10 minutes. 



Place the section on the slide, remove the surplus oil by 

 pressing over it several layers of tissue paper, and mount in xylol 

 balsam. 



Filter paper is avoided because it is liable to shed lint on the 

 section. Filter paper may be used over one thickness of tissue 

 paper. 



The above procedure should give a permanent isolated stain 

 for neuroglia fibers. Nuclei, neuroglia fibers, and fibrin should 

 stand out sharply in a clear blue. Everything else is decolorized 

 or appears a pale yellowish-gray tint. 



If the ferric chloride bath is omitted and the sections trans- 

 ferred directly from water (12) to 95 per cent, alcohol and 

 cleared and mounted (14), then, in addition to nuclei, neuroglia 

 fibers, and fibrin staining blue, the axis cylinders and nerve cells 

 stain a pale pink, and the ordinary connective tissue a deep pink. 

 Without the ferric chloride, however, the blue is not permanent, 

 and for the study of neuroglia it is better if other structures are 

 decolorized. 



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