BIELSCHO WSK Y 487 



central nervous organs, thoroughly fixed in 15 to 20 per cent, 

 formalin, are washed for some hours in running tap-water and 

 then cut by means of a COg freezing microtome. The sections 

 are collected in distilled water, thoroughly washed therein and 

 passed into a 2 or 3 per cent, solution of silver nitrate where they 

 are left for twenty-four hours in a dark place, and at room tempera- 

 ture. The sections can also be passed first into pure pyridine 

 for tzcenty-four to forty-eight hours, washed in many changes of 

 distilled water until the pyridine has been completely eliminated 

 and then transferred into 2 or 3 per cent, silver nitrate as above. 



The pyridine bath is optional and has the advantage of ensuring 

 a sharper stain of axis-cylinders whilst neuroglia, which is more 

 or less coloured when the pyridine bath is dispensed with, remains 

 unstained. Also connective tissue and nuclei are generally very 

 faintly stained after the pyridine treatment. Intracellular 

 neurofibrils, however, are not always so well shown as by the 

 direct passage of sections into the silver nitrate solution. 



Before proceeding further, one should prepare the Bielschowsky 

 ammoniacal silver nitrate-and-oxide bath as follows : Pour 

 5 c.c. of a 20 per cent, solution of silver nitrate into a measuring 

 cylinder and add to it first 5 drops of a 40 per cent, solution of 

 NaOH, and then ammonia, drop by drop, until the brown pre- 

 cipitate formed disappears ; dilute to 25 c.c. with distilled water, 

 and filter through paper washed with the same water. 



For staining, take sections one by one from the silver nitrate 

 bath, quickly wash them in distilled water for not more than 

 a few seconds, and transfer them into the ammoniacal silver 

 bath. Here they remain for about ten minutes when they become 

 yellowish-brown and should be, once more, quickly washed in 

 distilled water and placed in 20 per cent, formalin prepared with 

 tap water. The reduction takes place immediately, and if 

 one works with a number of sections it is advisable to re-transfer 

 them into a fresh bath of 20 per cent, formalin. 



At the end of half an hour and even less, the reduction can 

 be considered as accomplished and sections can be washed in 

 distilled water and toned with a diluted (0-2 per cent.) solution 

 of gold chloride. This may be slightly acidified with acetic 

 acid if one wishes to obtain a faintly purple background, or 

 neutralised with a few drops of a diluted solution of sodium 

 or lithium carbonate if one prefers greyish-white backgrounds. 

 Instead of gold chloride one can use a slightly acid solution of 

 chloroplatinic acid. After toning there remains only once more 

 to wash the sections in distilled water, and to pass them for a 

 few minutes into a 5 per cent, solution of sodium hyposulphite, 

 or any diluted fixing bath for photographic plates. Wash again, 

 dehydrate in alcohols of increasing strength up to 95 per cent., 

 clear in carbol-xylol, and mount in balsam. 



