20 CYTOLOGICAL TECHNIQUE 



'denatured'.^' ^' ^^^' ^^^ The most obvious change is loss of solu- 

 bihty with resultant coagulation. 



When a denaturing fixative is added to a solution of albumin, 

 coagulation usually follov^s so quickly that it appears to be 

 instantaneous. In fact, however, denaturation can be shown to 

 proceed by stages. The first eff'ect is an increase in reactivity; 

 there then follows a flocculation or aggregation into minute 

 particles, which are soluble in weak acids and alkalis; finally the 

 flocculi join into a clot, soluble only by reagents that cause 

 proteolysis. Flocculation and clot-formation are both included 

 in the term coagulation. Throughout all these changes the back- 

 bone of the protein remains unaltered. Some authors mean by 

 denaturation almost any change in a protein that increases 

 reactivity but does not destroy the backbone. ^^^ In this book the 

 term will be used in a more limited sense to indicate a marked 

 change involving the relation of the protein with the surrounding 

 water. This change results in coagulation. 



The chief hydrophil groups of proteins are the -NHg and 



O 



-C\ groups of basic and acidic amino-acids, the -OH group 



I 

 of tyrosine, and the C=0 groups of the backbone. Whether a 



particular protein be dispersed as a sol or held together by cross- 

 links to form a gel, it will ordinarily be related to water molecules 

 by such groups as these. When a protein is denatured, the rela- 

 tions with water are somehow disturbed. It may be supposed that 

 ethanol, a dehydrating agent, competes with the protein for the 

 water, and that the active groups, now no longer associated with 

 water, are more free to make fresh bonds with one another. The 

 formation of fresh bonds, so close as to exclude much of the 

 water that formerly lay between protein chains, would cause 

 coagulation. Interfaces between 'dry' protein and the surround- 

 ing fluid would thus be formed; and since these would scatter 

 light, the transparent sol or gel would change into a white 

 coagulum. 



Various reactive groups that were present in the unaltered 



