41 



slightly with hydrochloric acid, (4 drops in each 100 c. c.), for a day and then 

 remove to 80% alcohol. It affords a bright red stain that is quite transparent. 



100. Paracarmine (Mayer). Formula: Carminic acid, 1 gram; alumi- 

 nium chlorid, 0.5 gram; calcium chlorid, 4 grams; 70% (67%) alcohol, 100 c. c. 

 Allow it to stand a day or so, shaking occasionally until the carminic acid has 

 quite dissolved, and then filter. 



This is an excellent carmine stain for in toto staining. The tissue may be 

 stained one to several days (1 week), then washed in 67% and 82% alcohols to 

 remove the excess of staining fluid. A red nuclear stain, more opaque than borax 

 carmine. It does not over-stain readily, and since it is an alcoholic solution (70%) 

 it is quite penetrating and may be allowed to act for a greater length of time, 

 being thus suited for staining in toto objects of considerable size. 



101. Hcl. carmine. Formula: Carmine, 2 grams; concentrated hydro- 

 chloric acid, 3 c. c.; 70% alcohol, 100 c. c. Boil gently for 15 to 20 minutes to 

 dissolve the carmine; cool and filter. 



This is a strong carmine stain, quite suitable for sections, especially such as 

 stain with difficulty. It may also be employed for staining in toto. Stain sec- 

 tions from alcohol or water for 5 to 15 minutes ; rinse away the superfluous stain 

 with 67% (70%) alcohol and differentiate for a few seconds to a minute with acid 

 alcohol (95% alcohol 100 c. c., concentrated hydrochloric acid 1/10 c. c.). Wash 

 away the acid alcohol with ordinary 95% alcohol. If a pure nuclear stain is not 

 desired the differentiation may be omitted. 



Picric acid ( 116) may be used as a counter stain, and in that case differentia- 

 tion is ordinarily not required. 



102. Alum Cochineal. Formula: Powdered cochineal, 75 grams; potas- 

 sium alum, 75 grams; distilled water, 1000 c. c. Boil the ingredients for half an 

 hour, or (better) macerate for a day or so, boiling up two or three times. Cool 

 and filter. Add to the filtrate distilled water to make up 1000 c. c. and a crystal 

 of thymol as a preservative. 



This is an excellent stain, particularly for in toto staining of embryos, giving a 

 purpler stain than the carmine stains given above. 



Stain objects, such as embryos, over night to 2 days or longer, depending on 

 size. Wash out with water, 2 to 6 hours and place successively in 50%, 67% and 

 82% alcohols several hours to a day in each. 



103. Mucicarmine. Formula: Carmine, 1. gram; alummium chlorid 

 (pure), 0.5 grams; distilled water, 2 c. c. Mix thoroughly together and heat over 

 a small flame for 2 minutes (in a test tube) until the mixture has become dark red. 

 Dissolve the whole in 100 c. c. 50% alcohol added gradually; after 24 hours 

 filter. For use, it may be diluted 5 or 10 times with water. 



Stain sections 10 minutes or longer in the diluted stain, rinse in water, dehy- 

 drate, clear, and mount in neutral balsam. 



If desired, the stain may be diluted with 50 or 67% alcohol instead of water. 



104. Gentian Violet. Formula: A concentrated solution in distilled 

 water. Stain (paraffin) sections from water for 5 to 10 minutes, rinse in water, 

 dehydrate and differentiate with 95% alcohol and complete the differentiation 

 with clove oil. When the differentiation is sufficient, clear with bergamot oil 



