418 NERVOUS SYSTEM- SPECIAL METHODS. 



bedded in paraffin. The sections, which must be rather thin 

 (3 to 6 /x), are brought through xylol, absolute and 95 per cent, 

 alcohols into distilled water and here washed. 



This is the crucial point of the method because, by washing, 

 ammonium molybdate becomes extracted from the sections, and the 

 success of the subsequent staining depends almost entirely on 

 carrying out the extraction up to the right point. I find that the 

 only way of ensuring this consists in proceeding by trials, which must 

 be repeated for every series of sections. Once the right amount of 

 washing has been decided upon, one can proceed to stain even many 

 slides at the same time by means of a 1 : 10,000 solution of thionine, 

 to be freshly prepared every time from a less diluted stock solution. 



The staining is a " progressive " one, and must be controlled 

 under the microscope. It generally takes about twenty minutes to 

 obtain it, at the end of which time the grey substance has a red- 

 purple tone whilst the white substance appears bluish. If the 

 staining is right the preparations can be quickly washed, dehydrated 

 and mounted. But if the neurofibrils are not quite sharply stained, 

 the preparations can be " differentiated " for another fifteen to 

 twenty minutes in the ammonium molybdate solution used for 

 mordanting the pieces, or for ten seconds in a diluted solution 

 (1 : 10 to 1 : 20) of " pink salt " (C. Erba, Milano). Preparations 

 last only a few months, but are sometimes of great interest. See 

 DA FANO, Zieglers Beitrdge, xliv, 1908, p. 495. 



Method IV, which is particularly useful for the demonstration of 

 neurofibrils in the cells of the cortex cerebri and cerebelli, differs 

 from Method III only in regard to a preliminary fixation of pieces 

 for twenty-four hours in a mixture of pyridine nitrate 10 grms., and 

 pyridine, 100 c.c. ; they are then transferred for another thirty-six 

 hours into pure pyridine, proceeding as in Method III. 



Method V may be used for the demonstration of both NissPs 

 substance and neurofibrils. Pieces are fixed in a saturated solution 

 of corrosive sublimate ; after a day they are treated for twenty-four 

 hours with distilled water to which a few drops of iodine tincture 

 have been added, then for two to three hours with pure distilled 

 water ; and lastly passed for forty-eight hours into pure pyridine, 

 this being changed at least once. The rest as in Method III. 



PARAVICINI (Boll. Mus. Z. Anat. Comp. Torino, xx, 1905, p, 1) 

 fixes and mordants in the dark, and differentiates after staining with 

 extremely weak hydrochloric acid. 



TOMASELLI (Ztschr. wiss. Mikr., xxiii, 1906, p. 421) fixes spinal 

 ganglia for six to seven hours in absolute alcohol 100 c.c. with four to 

 five drops of ammonia, and then transfers them for two days into pure 



