224 APPENDIX 



time for the entire process on ordinary tissue should ex- 

 tend over two or three days. 



3. Staining : If the specimen is for temporary observa- 

 tion, the stains magenta or methylene blue may be used; 

 but for permanent mounts it is better to use alcoholic stains, 

 of which borax carmine (Grenacher's) is perhaps the most 

 desirable. The object should be placed in the stain from 

 sixty per cent alcohol, and should remain in the stain 

 until saturated to the degree of color desired. The length 

 of time required for this will have to be determined by 

 trial and depends upon the size of the spcimen, as well as 

 its penetrability. After staining, the object should be run 

 up to ninety per cent, and then to absolute alcohol, as di- 

 rected in 2, on hardening. Alcohol is useful for removing 

 an excess of stain, used with hydrochloric acid (3 to 6 drops 

 to 100 c.c of seventy per cent, alcohol) it will give a better 

 result from staining with borax carmine. The stained ob- 

 ject should be left in the acid alcohol for a time varying 

 from one-half hour to three hours.. This staining may be 

 done if desired after the sections have been cut and mounted. 



4. Clearing: After staining, the alcohol must be re- 

 moved by oil of cloves or turpentine. A short time only will 

 be necessary for this, and the specimen is then ready for 

 imbedding. 



5. Imbedding: For this purpose paraffin, melting point 

 50° C, should be melted and kept in a water bath at not 

 above 60° C. A softer paraffin will be found better some- 

 times, especially in cold weather. Drop the specimen to be 

 imbedded in this paraffin, and allow it to remain until it 

 is thoroughly permeated with paraffin. The length of time 

 varies from thirty minutes to an hour. 



