CHAPTER 2, 



FIXATION PROCEDURES 



Almost any tissue which is to be subjected to a cytochemical 

 procedure must first be fixed. The ideal result of fixation would 

 be to have an exact replica of the living tissue with each of the 

 molecules present in the original specimen in its original place, 

 and with no foreign molecules present. This is clearly an im- 

 possible ideal to achieve. In general, fixation can succeed only 

 in maintaining the large molecules, such as proteins, nucleic 

 acid, and carbohydrates, in approximately their original posi- 

 tions. In the case of the proteins, a considerable proportion of 

 the molecules are irreversibly changed. Indeed, if fixation is to 

 produce a specimen which will be satisfactory f6r any length 

 of time, the fixing agent used must effect two phenomena: it 

 must precipitate proteins and it must directly or indirectly 

 greatly restrict enzyme activity. It is not necessary that the 

 fixative should denature the proteins. For example, when a 

 heavy metal is the basis of a fixative, it precipitates the pro- 

 teins effectively but does not necessarily denature them. Simi- 

 larly, it is not necessarily the case that an enzyme should be 

 directly inactivated; it may be sufficient that it should be 

 rendered indiffusible by being captured by a matrix of another 

 protein. But if the enzyme systems of a cell are not, by one 

 method or another, prevented from displaying their normal 

 activity, the proteolytic and other hydrolytic enzymes rapidly 

 destroy the integrity of the tissue. 



Fixation is commonly carried out by the action of chemical 

 agents alone. Occasionally freeze-drying is used, but in the 

 past this has been somewhat uncommon. 



A major problem in the choice of fixative is presented by the 

 fact that, while a considerable amount of change must be 

 brought about in the specimen to render it insoluble in water 

 and to prevent enzyme activity, it is, nevertheless, necessary 

 that the group which is to be studied cytochemically shall not 



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