The finer Structure of the Nerve Cells of invertebrates. 17 



These results were most frequently obtained, when about 20 tlrops 

 of methylen blue had been added to the mixture of salt solution and 

 body fluid. 



2. Methods of Fixing and Staining. 



The following fixing agents were tried: chromic acid, Hermann's 

 fluid, Niessing's solutions, saturated solution of corrosive sublimate, 

 acetic sublimate and Flemming's solution. 



So far as my experience is concerned, Flemming's solution fixes 

 the nervous tissue of Invertebrates, in a manner superior to that of 

 all others; although Niessing's 2. solution (see 27) and Hermann's 

 fluid produce excellent results. 



A saturated solution of corrosive sublimate made up in a normal 

 salt solution, proved next to Flemming's solution the most useful 

 fixing agent. When using this solution, however, the attainment of 

 good results seemed in special instances (nerve cells of Crustaceans 

 and worms) to depend, as suggested by Heidenhain (see 15), upon 

 the use of bergamot oil for a clearing agent rather than xylol. 



Acetic sublimate, although highly recommended by Binet (1) 

 and others for the nervous tissue of Invertebrates, has in my ex- 

 perience proved a worthless fixing agent for such material. In no 

 case have I been able to obtain material fixed by it which has not 

 been considerably shrunken. The same may be said for tissues fixed 

 in chromic acid ^). 



For tissues fixed in Flemming's solution , the following stains 

 were chiefly used: Heidenhain's iron-alum haematoxylin method, 

 iron-alum and Delafield's haematoxylin without after-differentiation ^), 

 a weak solution of Delafield's haematoxylin as used by Flemming 

 (see 11), and a combination stain of safranin and light green which 

 will be described below. 



For tissues fixed in sublimate, a large number of stains were 

 tried. Among these, in addition to the haematoxylin stains mentioned 

 above, two combination or double stains proved most valuable. 



By far the best of these double stains was a combination of 

 methylen blue and eosin, used after a modification of Mann's method 

 (Rawitz, Leitfaden für histol. Untersuchungen, 2. Aufl., p. 70). 



1) After fixing, all material was gradually run through the alcohols, 

 cleared in xylol or bergamot oil, and embedded in paraffine. 

 3) After Flemming. 



Zool. Jahrb. XI Abth. f. Morph. 2 



