I 



1917I BRIEFER ARTICLES 411 



solution in 100 cc. of water are sufficient. The iron-alum solution is 



the 



After 



a short time, 15-30 minutes or even less being sufficient, the material 

 is washed thoroughly with distilled water from the wash bottle. A weak 

 haematoxylin stain is then applied slowly to the material, and repeated 

 f observations of specimen mounts under the microscope will determine 



when the staining is complete. A 0.1 per cent haematoxylin stain is 

 strong enough, and 30 minutes or less time is long enough for it to act. 



When 



distilled, 



destaining with the o . 1 per cent iron-alum solutions. This latter step 

 is the one requiring the greatest care, since it can be overdone most 

 easily; the light haematoxylin stain used in the method is easily lost 

 by prolonged treatment with even a very weak iron-alum solution. One 

 application of the destaining solution is generally sufficient if care be 

 taken that the material is thoroughly saturated with the solution. As 



1 



usual when using the iron-alum solution for differentiating a stain, the 

 process should be closely observed under the microscope. It is to be 

 understood that no directions concerning the time limits can be given 

 for the use of either the staining or differentiating solutions, since success 

 depends upon the proper balancing of these two processes. When 

 the stain has been properly differentiated, the material should be thor- 

 oughly washed with distilled water from the wash bottle. At any 

 stage of the process, should it be desired to allow the material to remain 

 covered by the solution being used, this can be accomplished by fitting 

 a short piece of rubber tubing over the stem of the funnel and using a 

 clamp to stop the flow of the solution. 



Dehydration is accomplished by the glycerine dehydration method 

 in general use. The transfer to the 5 per cent glycerine solution is made 

 by puncturing the filter paper and washing with the glycerine solution 

 into an open vessel, such as a Petri dish. The minimum amount of 

 glycerine should be used, since it is difficult to remove. After a few 

 days' exposure to evaporation, the glycerine solution is concentrated. 

 The material is again poured into the filter paper in the funnel and 

 complete dehydration accomplished by washing all the glycerine out 

 with 95 per cent alcohol, followed by absolute alcohol to complete 

 dehydration. There should be no doubt about the thoroughness of 

 this part of the process, since complete dehydration is essential to 

 success. When completely dehydrated the material is transferred 

 quickly to a 10 per cent Venetian turpentine solution and placed in a 



