CHAPTER II 



TECHNIQUE OF FIXING, IMBEDDING, AND STAINING 



CHAMMKKI.AIN. C. J. Methods in Plant Histology. (2d ed.) 262 pp. 



1905. 



LEE, A. B. Microtomist's Vade Mecum. (6th ed.) 538 pp. 1905. 

 ZIMMKR.MAN. A. (Transl. by J. E. Humphrey.) Botanical Microtechnique. 



296pp. 1893. 



I. FIXING 



The purpose of a fixing agent is to kill and fix, or render per- 

 manent, the structural relations of the cell and the associations of 

 cells in tissues. The finer protoplasmic structures are readily dis- 

 organized and lost for study, if not carefully fixed by special agents. 

 Moreover, adequate fixing is necessary in order to prepare tissues 

 to show properly the differential effects which may be gained by 

 staining. It is well, therefore, to fix by one or more of the best 

 methods such material as may be valuable for minute microscopic 

 study. This, however, in no way precludes the desirability of study- 

 ing material in a living condition also, whenever that is possible. 



The material to be preserved should be plunged into the fixing 

 solution in a condition as fresh as possible, so that no changes may 

 occur subsequent to removal from the natural substratum. Great 

 haste is often necessary with delicate fungi in order to avoid dry- 

 ing out. It is well always to use an abundance of the fixing liquid. 

 \Yith osmic and chromic acids one should often employ as much 

 as fifty times the quantity of the liquid as of the material, while 

 with alcohol and formalin, fully three times as much liquid as 

 material. In all cases, the object, if large, should be cut into pieces 

 as small as practicable, in j-inch cubes or less. 



Fixing methods and fixing agents are numerous, and the method 

 or agent to be selected will depend upon the kind of study for 

 which the material is desired. One method will be applicable when 

 histological differentiation is the chief end sought, and when a study 



