FATTY SUBSTANCES 285 



in (B) for three days at 37° C. ; (2) rinsed in distilled water ; 

 (3) mounted on a slide ; (4) blotted dry with filter paper ; 

 (5) treated with a few drops of (C) ; and (6) covered with a 

 cover-glass. 



665. Notes on Technique. (1) The acid mixture should be 

 made by putting the glacial acetic acid in a flask and slowly adding 

 the sulphuric acid with careful shaking, the flask being kept cool 

 by immersion in cold water. (2) Both acids must be of " analytical 

 reagent " standard, and the sulphuric at least 98 per cent. H2SO4. 

 After the acid mixture has been added to a section a few bubbles 

 usually appear, but do not interfere with the appreciation of 

 colour. If the acids are impure, however, bubbles may appear in 

 large numbers and cause serious difficulty. The acid mixture is 

 hygroscopic and the bottle containing it must therefore be kept 

 stoppered when the mixture is not in use. If due care is exercised 

 the acid shows no sign of deterioration even after a year's regular 

 use. (3) If the iron alum solution is heated during preparation 

 basic iron sulphate is precipitated ; this cannot be redissolved. 

 Both 10 per cent, and 2-5 per cent, solutions keep well. The 

 2-5 per cent, solution is discarded after being used once. The object 

 of preparing a 10 per cent, solution is to reduce labour when large 

 volumes of the 2-5 per cent, solution are being used. (4) Gelatine 

 embedding, advised by Schultz (1925), is unnecessary. A film of 

 egg albumen on the slide keeps the section flat during the perform- 

 ance of the test. The test is applicable also to urinary deposits 

 and sputa, which are dried on a slide, fixed in formol, and then 

 placed in the iron alum solution. (5) At the start of a test a dry 

 glass rod should be inserted in the acid bottle and returned to it 

 immediately after it has been used to add acid to a section. If the 

 rod is left exposed to the air the film of acid on the rod will be 

 rapidly diluted by atmospheric moisture. At the end of a test 

 the acid bottle should be stoppered and the glass rod washed in 

 water and dried. (6) The blue-green colour appears within a few 

 seconds after the acid mixture has been added and becomes 

 stronger during the next few minutes. It becomes dirty brown 

 within half an hour and sections must therefore be examined 

 without delay. Permanent preparations are unobtainable. Fatty 

 substances giving a negative result appear brown. (7) The 

 operator may check his reagents and technique by testing sections 

 of a suprarenal gland from a guinea-pig or rabbit killed in good 

 health ; the fatty substances in the cortex give a strongly positive 

 result, even in sections that have been kept in formol for months, 

 (8) Whenever a section gives a negative result, further sections 

 should always be examined to exclude technical errors. Sections 

 of a tissue known to be Schultz-positive should be tested at the 

 same time. 



