F02 



FIXATIVES 



187 



potassium dichromate, each in its own 

 solution, are used successively in the 

 course of fixation. These have been classi- 

 fied for the sake of simplicity as thouj^h 

 both reagents were included in the same 

 solution. The last case of ambiguity which 

 occurs is that in which a complex salt, 

 involving two primary fixative agents as 



one compound, is utihzed. There are 

 several formulas, for example, in which 

 copper dichromate is specified as an in- 

 gredient. In these cases the formulas have 

 been classified as though they were com- 

 posed of a mixture of copper and dichro- 

 mate rather than as prepared from the 

 compound. 



F 02 General Observations 



The great confusion and diversity of 

 opinion which occurs in the literature as 

 to what constitutes a desirable quality for 

 a fixative ingredient, is probably due to a 

 similar disagreement as to what consti- 

 tutes a desirable quality in a fixative mix- 

 ture. In general, those who study the form 

 of small invertebrates require first that a 

 fixative should not interfere with the 

 recognition of the object after fixation, 

 that is, that its general shape shall not be 

 distorted by the fluid applied to it. Cytol- 

 ogists, histologists, and pathologists, on 

 the contrary, require as a first considera- 

 tion that the inner structure of the cell 

 constituent shall remain unaltered, or 

 shall at least present on microscopic ex- 

 amination only those features which are 

 thought to have been present in the 

 original hving cell. It is obvious that the 

 first group of workers are dependent for 

 their effect largely upon the osmotic 

 pressure shown by the material or, alter- 

 natively, they require a solution that so 

 rapidly and thoroughly hardens the outer 

 coat that it will not become distorted by 

 the passage of solutions through it. Thus 

 Young (1935) gives directions for adjust- 

 ing the osmotic pressure of solutions by 

 the addition of sodium chloride, and Gray 

 1933 (11360, 53:13) pointed out that the 

 addition of sodium sulfate to potassium 

 dichromate to give the solution of Miiller 

 improved the osmotic performance of the 

 mixture. Heat is considered by many to 

 be a desirable characteristic; Pantin 

 (1946, 8) recommended heating fixatives 

 for small marine invertebrate larvae. Heat 

 should also possibly be apphed to fixation 

 of internal constituents of cells, since 

 Thomas and Morris 1925 (10996, 48:501) 

 showed that the dichromates do not pre- 

 cipitate albumin unless they are heated 



either at the same time, or immediately 

 after, the apphcation of the reagent. The 

 principal opponents of the view that the 

 osmotic pressure of the solution is of 

 primary importance are Baker and Craw- 

 ford {test. Langeron 1942, 362) who showed 

 that solutions with high osmotic pressure 

 do not of necessity cause distortion; and 

 Hirsch and Jacobs 1926 (1820, 18:7) who 

 denied emphatically that any improve- 

 ment in fixing qualities could be found 

 by the addition of sodium chloride to 

 fixative solutions. 



Those who desire to fix only the con- 

 stituents of the living cell have been them- 

 selves sharply divided into two schools of 

 thought. The first of these schools con- 

 sidered that the precipitating, or coagu- 

 lating powers of the reagents employed 

 were of primary importance. The second 

 considered penetration and pH to be the 

 most critical characteristic of a fixing 

 fluid. The school which depended upon 

 coagulation for fixation appears nowadays 

 to be so thoroughly discredited that one 

 can only refer to the summary of their 

 work given in B5hm and Oppel (1907, 14). 

 Here will be found a complex classification 

 of fixatives based entirely upon the mate- 

 rials which were precipitated and rendered 

 insoluble by the apphcation of various 

 reagents. There is, indeed, a fatal objec- 

 tion to this work which was pointed out 

 by Bolles Lee (1905, p. 22). This is that 

 the more organic dead materials are pre- 

 cipitated by the solution employed, the 

 greater will be the artifacts produced. The 

 case for cytologists and histologists is 

 probably best given by Cretin (1925, 

 LeMans) who considers that for neutral 

 solutions, the total concentration of salts 

 is important, but that for any solution 

 containing acid, the pH must be carefully 



