108 CABMINE AND COCHINEAL STAINS. 



precipitates in the cavities of bulky objects which cannot be 

 removed by washing out. And it must be remembered that 

 the fluid is alkaline, and therefore not suitable for delicate 

 cytological work. I believe that some otherwise excellent 

 cytological work has been vitiated by over-confidence in this 

 reagent, and will have to be done over again. 



170. Alcoholic Hydrochloric Acid Carmine. In view of the above- 

 mentioned defects of borax-carmine defective penetration, and a tendency 

 to form insoluble granular precipitates in the interior of objects it is 

 desirable to possess a powerful staining medium more highly alcoholised and 

 of acid reaction. Hydrochloric acid carmine possesses these qualities, and 

 may be useful in the case of objects for which Mayer's paracarmine may 

 not suffice. It may, for instance, be frequently useful in work on Arthropoda, 

 especially the marine forms. 



GBENACHEE'S receipt (Arch. f. tnik. Anat., xvi, 1879, p. 468) will be 

 found extremely troublesome by those who are not expert at neutralising. 

 The following method, due to PAUL MAYEE (quoted from GASSINGS 

 Manuals per la Technica moderna del Microscopic, first ed., p. 46), is easy 

 and gives excellent results. Take 100 grms. of alcohol (either absolute or of 

 any weaker grade), 1 or 2 drops of HC1, and an excess of carmine, and boil 

 until you get a clear solution, taking care that there remain an excess of 

 carmine. This ought to give a nuclear stain, without the aid of HC1 for 

 washing out. If overstaining or diffusion should occur, wash out with 

 alcohol, very slightly acidulated with HC1. 



BEASS (Zeit. f. wiss. Mik., ii, 1885, p. 303) takes 100 c.c. of 70 per cent, 

 alcohol, 15 drops of HC1, and an excess of carmine. An old formula of 

 PAUL MAYEE'S (M. T. Zool. Stat. Neapel, iv, 1883, p. 521 ; Journ. Roy. 

 Mic. Soc. [N. S.], iv, 1884, p. 317), which gives a more powerful stain than 

 the preceding, is as follows : 



Four grms. carmine are dissolved in 100 c.c. of 80 per cent, alcohol with the 

 addition of 30 drops of concentrated pure hydrochloric acid, and heated for 

 about half an hour in the water-bath ; the solution is filtered whilst still 

 hot, and the superfluous acid is carefully removed by the addition of caustic 

 ammonia, added until the carmine begins to be deposited. This solution 

 stains very rapidly (embryos of lobsters are stained in about a minute) and 

 intensely, though diffusely ; the preparations must be washed out with HC1 

 alcohol if a nuclear stain is required. 



The heating of alcohol of so high a grade as 80 per cent, being trouble- 

 some, not to say dangerous, the process may be modified by dissolving the 

 carmine in 15 c.c. of water acidulated with the HC1, adding 95 c.c. of 85 per 

 cent, alcohol, and then neutralising with ammonia. 



If it be desired to dilute any of these solutions it should be done with 

 alcohol, not water, and alcohol should be taken for washing out. 



