206 GUIDANCE IN MAKING PERMANENT PREPARATIONS. 



The time an object should be treated with stain varies with 

 the stain and the size of the object. Alum-carmine should be 

 used from six to twenty hours, according to circumstances. 

 Borax-carmine should be used from five minutes to half an hour. 

 Aceto-carmine, used for killing and staining, acts very rapidly. 

 Delafield's hematoxylin (a dark wine-colored solution in water) 

 requires ten minutes to half an hour. In all these cases, exam- 

 ination of the objects themselves is the only means of deciding 

 when staining is sufficient. It is usually best to slightly over- 

 stain and then to bleach out, as certain parts of the protoplas- 

 mic structure will retain the stain better than others and thus 

 better differentiation will be secured. After staining, bring 

 the tissues gradually into 70 percent alcohol, and then treat 

 with acidulated alcohol to remove excess of stain. After this, 

 every trace of the acid must be removed by washing in clean 

 alcohol, or the tissues will continue to bleach after they are 

 mounted. The specimen is now ready for final dehydration. 

 In damp climates, as at the seashore, your stronger alcohols 

 must be kept closely covered all of the time or they will take 

 water from the atmosphere and be unfit for the purpose. Run 

 through 80 percent, 95 percent, and 100 percent alcohol, thus 

 completing dehydration. Every trace of water must be removed 

 and then kept out. 



Clearing and Mounting. 1 From absolute alcohol, place ob- 

 jects in some clearing fluid (clove oil, cedar oil, or xylol) and leave 

 till they have a clear, translucent appearance, after which place 

 on a dean slide, with some Canada balsam or dammar, and cover 

 with a cover-glass. 



If the object turns cloudy or milky when placed in the clean- 

 Ing fluid, it is evidence that all of the water has not been removed, 

 and it should be returned to absolute alcohol for complete dehy- 

 dration. Tissues left in the clove oil or xylol for any great 

 length of time will become hard and brittle. In case tissues 

 in the process of preparation must necessarily be left untreated 



1 Specimens may be successfully mounted in upural from 95 percent 

 alcohol. This avoids labor of dehydration and clearing and gives perma- 

 nent mounts. 



