STUDY OF THE ACTION OF FIXATIVES 81 



some material, such as rat liver, a prolonged period of fixation 

 does not seem to result in the production of artefacts. But as 

 Cain and others have emphasized, artefacts probably due to oxi- 

 dation arise rather readily in some tissues; I have found this 

 to be the case with a number of tumor tissues. 



Cain has suggested that a better technique is to use very small 

 pieces of tissue which are placed, without previous fixation, in a 

 mixture containing mercury and reduced fuchsin. With the tis- 

 sues which I have studied, fixation under these circumstances 

 tends to be very poor. But in any case there is always, with tech- 

 niques of this type in which the colour due to a cytochemical re- 

 action is produced in a block of tissue, the fundamental objection 

 that it is impossible to study the extent to which the results are 

 invalidated by diffusion phenomena. It is my view that a cyto- 

 chemical technique which does not permit the evaluation of diffu- 

 sion artefacts is useless. 



After a preliminary survey of a tissue made by the procedure 

 given above, the other points mentioned in the introduction to 

 this chapter may then be studied as seems appropriate. 



Study of the Action of Fixatives 



Where the substance which is being studied can safely be 

 passed through an infiltration with wax, freeze-drying is the best 

 method of fixation. But, for many tissue aldehydes, organic 

 solvents must be avoided since they dissolve fatty aldehyde ma- 

 terial from the tissue. 



When freeze-drying cannot be employed it is necessary to use 

 a variety of fixatives to investigate the degree to which the dis- 

 tribution of aldehyde is produced as a fixation artefact, as dis- 

 cussed in Chapter 2. It is possible to use acid, neutral, and alka- 

 line fixatives, reducing fixatives, and preprecipitation with strong 

 salt solutions in the study of fixation artefacts. There are, of 

 course, limitations encountered in the use of this variety of fixa- 

 tives with particular types of aldehyde. Thus an acid fixative 

 is likely to liberate aldehyde from acetals, after which acetal 

 adehyde and free aldehyde are, of course, indistinguishable. 



As an example of the use of this method, we may consider 

 studies which have been made of the fatty aldehydes of liver. 

 When this material is placed in fixatives containing mixtures of 

 formaldehyde, acetic acid, mercuric chloride, pyridine, and tri- 



