550 POWDERED DRUGS 



differentiating pectin compounds. The protoplast and lignified walls are stained a 

 bright blue, while pectin compounds are stained a violet blue. Cells containing 

 tannin will accumulate methylene-blue from very dilute solutions. The sections of 

 living tissues are placed in a solution of i part of the stain in 500,000 parts of filtered 

 rain-water. The cells containing tannin soon take on a distinct blue color, and, 

 later, a deep blue precipitate is formed in them. The gelatinous sheaths of live 

 conjugates may be stained by dilute aqueous solutions of methylene-blue without 

 injury to the living organism. A o.ooi per cent, solution of methylene-blue in 

 water will stain the living nuclei of diatoms and other simple organisms. The cen- 

 tral body of the Cyanophyceae may be stained by the above dilute solution if, after 

 24 hours' treatment, the stain is strengthened to a o.i per cent, solution. Methy- 

 lene-blue and carmine form a good differential stain for bacteria occurring in 

 sections of tissues. 



Methylene-blue and Carbol-fuchsin. This double staining method is used in 

 the differentiation of Bacillus tuberculosis. The material first coughed up from the 

 lungs by the patient on waking in the morning should be expectorated into a wide- 

 mouthed bottle or covered jar. The person who is to make the examination 

 should afterward pour this out into a shallow glass dish. This should be placed on 

 a dead-black background, and one of the small, yellowish, lenticular bodies which 

 usually occur in tuberculous sputum should be removed and placed on a cover- 

 glass. A second cover-glass should be placed over this ; then press the cover-glasses 

 gently between the thumb and forefinger, and rub to and fro until the material is 

 spread out in a thin film on the cover-glasses. Then slide the cover-glasses apart, 

 and allow them to dry in the open air. When dry, hold them with a pair of for- 

 ceps and pass them 3 times through the flame of the Bunsen burner or alcohol 

 lamp. (The film should not be allowed to turn brown, else the preparation will 

 be ruined.) Next pour over them carbol-fuchsin prepared by rubbing i Gm. of 

 fuchsin with 100 mils of a 5 per cent, aqueous solution of carbolic acid, with the 

 gradual addition of 10 mils of alcohol. Hold the cover-glasses over a flame with 

 forceps until vapor begins to arise from the surface of the stain. Then hold away 

 from the flame, except in intervals of gentle heating, by which they are kept warm 

 for a minute or two. They are next washed in water and decolorized by being 

 moved about in a 25 per cent, solution of nitric or sulphuric acid. When the pre- 

 viously deep-red color has changed to a greenish tint, the preparation is washed in 

 60 per cent, alcohol to remove the color set free by the acid. If any red color still 

 remains, the preparation should be rinsed in water and again treated with the acid- 

 bath. By the above process the fuchsin has been removed from everything but 

 the tubercle bacilli. The double staining is accomplished by now pouring over the 

 preparation a mixture of 3 parts of water with I part of a concentrated alcoholic 

 solution of methylene-blue. After a few minutes the methylene-blue is washed 

 off with water, and the preparation is allowed to dry; when dry, it may be mounted 

 in Canada balsam. Other bacteria than the tubercle bacilli are decolorized by the 

 acid-bath, and are subsequently stained blue by the methylene-blue. 



Methyl-green. An aqueous solution of this stain serves well to differentiate 

 the nucleus of cells containing aleurone grains. Sections through vascular bundles 

 which have been treated for some hours with alcohol borax-carmine, and then for 

 a^short time with methyl-green, have the protoplasmic cell-contents stained red, 

 the lignified walls of the tracheal tubes green, and the walls of the primary phloem 

 portion green. 



Methyl -green and Acetic Acid. Methyl-green is dissolved in a 2 per cent, 

 solution of acetic acid until the solution has a blue-green color. The nuclei of 

 fresh material teased out in this become instantly fixed and stained. It is very use- 

 ful for a preliminary examination of dividing nuclei. 



