FIXATIVE MIXTURES 149 



of new embedding media is likely to result in less reliance being 

 placed on coagulant fixatives. 



It follows from what was said in chapter 6 that fixative mixtures 

 intended for the study of cytoplasmic inclusions will contain 

 formaldehyde, osmium tetroxide, or unacidified potassium 

 dichromate, or more than one of these. Acetic acid is omitted or 

 reduced to a minute amount. These generalizations apply to Alt- 

 mann, Aoyama, Benda, Bensley's AOB, Champy, Helly, Lewitsky, 

 Mann, Orth, Regaud, and Zenker without acetic. Of these, only 

 Benda and AOB contain acetic acid, at great dilution (less than 

 0-2%). It must be emphasized once more that the omission or 

 great reduction of acetic acid is not necessarily connected with pH. 

 Thus Lewitsky differs from Flemming's strong fluid solely in the 

 absence of acetic acid from the former: yet the two have the same 

 pH.^^^ Lewitsky's fluid was designed specifically to fix mito- 

 chondria, while Flemming (with full acetic) destroys them. This 

 shows the specific destructive (or at any rate non-fixative) effect of 

 acetic acid on mitochondria. This effect, as we have seen (p. 138), 

 is not shared by hydrochloric acid. Acetic or any other acid, unless 

 very dilute, will transform potassium dichromate into a coagulant 

 fixative and in so doing abolish its ability to fix mitochondria. 



Coagulants are often omitted from mixtures used for the study 

 of cytoplasmic inclusions. This applies, for instance, to Altmann, 

 AOB, Orth, and Regaud. Easy infiltration with paraffin is sacri- 

 ficed for the sake of homogeneous fixation of the ground cyto- 

 plasm. It is particularly difficult to obtain good paraffin sections of 

 tissues fixed in Altmann. 



Each component of a fixative mixture should so far as possible 

 compensate for a defect in another. Thus ethanol shrinks tissues 

 strongly and does not fix chromatin: acetic acid compensates for 

 both these defects. Acetic acid does not fix cytoplasm or nuclear 

 sap: ethanol fixes both (though the former indifferently). When 

 ethanol and acetic acid are mixed to make Clarke's fluid, an 

 excellent fixative for routine histolog}^ and chromosome studies is 

 produced. 



The properties of Bouin's fluid can be analysed in a similar way. 

 Formaldehyde fixes cytoplasm and nuclear sap, but hardens 

 tissues unduly, prevents paraffin from penetrating easily, makes 

 cytoplasm basiphil (so that acid dyes do not work well), and fixes 

 definitive chromosomes poorly. Picric acid compensates for most of 

 these defects. It leaves tissues soft, coagulates cytoplasm in such a 



