4 INTRODUCTORY. 



action of the alcohol being continuously prolonged, the minute structure of 

 tissues is sometimes considerably altered by it ; they become overhard and 

 shrink, and become brittle, and their capacity for taking stains well becomes 

 seriously diminished. KULTSCHITZKY (Zeit. f. wiss. Mik., iv, 3, 1887, 

 p. 349) has proposed to remedy this by putting up objects, after fixation and 

 washing out with alcohol, in ether, xylol, or toluol. FLEMMING has lately 

 (Arch. f. tnik. Anat., xxxvii, 1891, p. 685) advised putting up objects after 

 fixation in a mixture of alcohol, glycerin, and water, in about equal parts, 

 pointing out that objects thus preserved may be at any moment either pre- 

 pared for sectioning by treatment with pure alcohol, or softened for dissec- 

 tion or teasing by a little soaking in water, and that they do not become so 

 hard and brittle as alcohol specimens, and retain their staining power much 

 better. After extensive experience of this plan, I can highly recommend it, 

 and would only further suggest that the action of the liquid seems to me to 

 be in many cases much improved by addition of a little acetic acid (say 0'5 

 to 0'75 per cent.). 



The water having been thus sufficiently removed, the 

 alcohol is in its turn removed from the tissues, and its place 

 taken by some anhydrous substance, generally an essential 

 oil, which is miscible with the material used for imbedding. 

 This operation is known as Clearing. It is very important 

 that the passage from the last alcohol to the clearing agent be 

 made gradual. This is effected by placing the clearing medium 

 under the alcohol. A. sufficient quantity of alcohol is placed 

 in a tube (a watch-glass will do, but tubes are generally 

 better), and then with a pipette a sufficient quantity of clear- 

 ing medium is introduced at the bottom of the alcohol. Or you 

 may first put the clearing medium into the tube, and then 

 carefully pour the alcohol on to the top of it. The two fluids 

 mingle but slowly. The objects to be cleared being now 

 quietly put into the supernatant alcohol, float at the surface 

 of separation of the two fluids, the exchange of fluids takes 

 place gradually, and the objects slowly sink down into the 

 lower layer. When they have sunk to the bottom, the alcohol 

 may be drawn off with a pipette, and the objects will be found 

 to be completely penetrated by the clearing medium. (It may 

 be noted here that this method of making the passage from 

 one fluid to another applies to all cases in which objects have 

 to be transferred from a lighter to a denser fluid for instance, 

 from alcohol, or from water to glycerine. It is a more exact 

 method than that of successive baths of mixture of alcohol 

 and clearing agent.) 



The objects are now imbedded. They are removed from 



