PIXINU AND HARDENING AGENTS. 57 



91. Nitrate of Copper (Gilson, from Gelderd, La Cellule, xxv, 

 1909, p. 12).— Nitrate of copper 200, formol 500, sea-water 200. Seven 

 parts of this solvition to be diluted with 100 of sea- water. For Crustacea. 



92. Acetate of tTranium (Schbnk, Mitth. Enibryol. Inst. Wten, 

 1882, p. 96 ; of. GiLSON, La Cellule, i, 1885, p. 141) has a mild fixing 

 action, and a high degree of penetration, and may be combined with 

 methyl green. 



Pkiedenthal {Sitzh. Ges. Nat. Freunde Berlin, 1907, p. 209) recom- 

 mends equal parts of saturated solution of the acetate and trichlor- 

 acetic acid of 50 per cent. 



93. Picric Acid. — Picric acid in aqueous solution should 

 be employed in the form of a strong solution whenever it 

 is desired to make sections or other preparations of tissues 

 with the elements in situ, as weak solutions macerate ; but 

 for dissociation preparations or the fixation of isolated cells, 

 weak solutions may be taken. Flemming finds that the 

 fixation of nuclear figures is equally good with strong or 

 weak solutions. The saturated solution is the one most 

 employed. (One part of picric acid dissolves in about 86 

 parts of water at 15° 0. ; in hot water it is very much more 

 soluble.) Objects should remain in it for from a few seconds 

 to twenty-four hours, according to their size. For Infusoria 

 one to at most two minutes will suflftcOj whilst objects of a 

 thickness of several millimetres require several hours. 



Picric acid should always be washed out ivith alcohol, that 

 of 70 per cent, being mostly indicated. Staining should be 

 performed by means of alcoholic solutions, or if with aqueous, 

 then with such as are themselves weak hardening agents, 

 such as heemalum, carmalum, methyl green. 



Washing out is facilitated by heat, the extraction being 

 about twice as rapid at 40° C. as at the normal temperature 

 (Pol). 



It has been found by Jblinek [Zeit. wiss. Mik., xi, 1894, 

 p. 242) that the extraction is greatly quickened by the 

 addition of a base to the wash-alcohol. He recommends 

 carbonate of lithia. A few drops of a saturated solution of 

 the salt in water are added to the alcohol ; a precipitate is 

 formed. The objects are put into the turbid alcohol, which 

 becomes clear and yellow in proportion as the picrin is 

 extracted. Further quantities of carbonate are added from 

 time to time until the colour has been entirely extracted. 



