6 INTRODUCTION 



cal side are far more easily underestimated than otherwise. As 

 examples of some of the problems which affect one from the bi- 

 ological point of view, may be mentioned fixation and diet. It 

 is doubtful whether any method of fixation other than freeze- 

 drying is really adequate for cytochemical purposes. Then, 

 as far as diet is concerned, it will be a source of amazement 

 to future generations to discover in how few of the papers on 

 cytochemistry the diet of the animals used is at all defined. 

 Yet the cytochemical pattern in organs such as the liver and 

 intestine is astonishingly dependent upon diet. As another ex- 

 ample, may be mentioned the work of Dr. H. Mugard (1953), 

 who has recently shown that the cytoplasm of the ciliate 

 Ophryoglena atra which has not recently fed is apparently free 

 from the enzyme alkaline phosphatase: yet within a few seconds 

 of the formation of a food vacuole a high concentration of phos- 

 phatase is present in the cytoplasm. For particular studies it 

 will no doubt be equally important to define the age of the 

 animal under investigation, the time of taking the specimen, 

 the state of hormone activity, and other biological variables, 

 before a generalisation on the cytochemical level may safely 

 be embarked upon. 



Before passing on to consider the fields in which I have played 

 some part in developing techniques, a number of techniques 

 will be considered critically, since by so doing it is possible 

 to see many of the major hazards which exist in the field of 

 cytochemistry, and which it has been my endeavour to avoid. 



Centrifugal Stratification, etc. 



In these techniques the common principle is, by centrifuga- 

 tion or other methods, to separate parts of cells which differ 

 from one another in their physical characteristics. The most 

 usual approach is to stratify a cell by centrifugation, to separate 

 the strata, and then examine the distribution of various sub- 

 stances in the different strata. In the hands of Holter and of 

 Shapiro, this work has provided valuable information about 

 the distribution of certain substances, particularly enzymes, in 

 large cells such as amoebae and echinoderm eggs. There are, 

 however, some very marked limitations to techniques of this 

 type. Their use is usually limited to large individual cells: 



