40 FIXING AND HARDENING AGENTS. 



Whitman finds one hour sufficient, and transfers to 50 per cent, 

 alcohol. 



A similar mixture, with the addition of 0-25 to 0-1 per cent, of 

 acetic acid, is recommended by BRASS for Protozoa ; and LAV- 

 DO WSKY has used for nuclei a mixture of 10 parts of 1 per cent, 

 chromic acid, 5 of 1 per cent, platinum chloride, and 100 of 5 per 

 cent, acetic acid. 



Whitman recommends for the hardening of pelagic fish ova a 

 stronger mixture (due, I believe, to Eisig), viz.- 



0-25 per cent, solution of platinum chloride . . 1 vol. 



1 per cent, solution of chromic acid . . 1 



The ova to remain in it one or two days (WHITMAN, Methods in 

 Micro. Anat., p. 153). 



Salts. 



50. Chromates. The chromates are amongst the oldest and best 

 tried of hardening agents. The bichromate of potash especially 

 was at one time universally employed for hardening all sorts of 

 tissues. 



FLEMMING (Arch. mik. Anat., xviii, 1880, p. 352) pointed out that 

 though it preserves cytoplasm well it causes chromatin to swell, and 

 therefore should not be employed for the study of nuclei. But, duly 

 corrected with acetic acid, it affords a correct and fine fixation of 

 nuclei ; whilst preserving hyaloplasm and its inclusions, secretions, 

 etc., much better than chromic acid. 



For an elaborate study of the action of chrome salts on nucleus and 

 cytoplasm, see BURCKHARDT, La Cellule, xii, 1897, p. 335. He finds 

 that the bichromates of sodium, ammonium, magnesium, strontium, 

 and zinc have the same destructive action on nuclei that the bichromate 

 of potassium has ; but that the bichromates of barium, calcium, and 

 copper have not. He concludes that acetic acid ought always to be added, 

 not only to ensure the correct fixation of nuclei, but also to enhance 

 penetration and the good preservation of cytoplasm. 



The following is recommended by him as a good combination for tin 1 

 fixation both of cytoplasm and nucleus : 



Bichromate of barium, 4 per cent, solution . . 60 vols. 

 Bichromate of potassium, 5 per cent, solution . . 30 ,, 

 Glacial acetic acid . . . . . . 5 ,, 



(Instead of the barium you may take. 4 per cent, solution of bichromate 

 of calcium, or 6 per cent, solution of bichromate of copper.) 



For the demonstration of the achromatic figure of cell division he 

 recommends 



Chromic acid, 1 per cent, solution . . . .60 vols. 



Bichromate of potassium, 5 per cent, solution . . 30 ,, 

 Glacial acetic acid . 5 ,, 



