REAGENTS AND PROCESSES 557 



running water may be used to wash out the fixative whether the latter has been 

 dissolved in alcohol or in water. 



Picro-aniline Blue. A double stain, which is very rapid in its action, is prepared 

 by adding aniline blue to a saturated solution of picric acid in 50 per cent, alcohol 

 until the solution has a blue-green color. By this treatment the unmodified cell-, 

 walls and the cell-contents are stained blue, while the lignified walls are stained 

 by the picric acid. 



Picro-nigrosin. A solution of nigrosin in a concentrated solution of picric acid 

 in water or 50 per cent, or 95 per cent, alcohol is a good fixative and stain for algae 

 and leucoplasts, and for double-staining modified and unmodified cell- walls. The 

 solution may, in some cases, need to act for 24 hours. The strong alcoholic 

 solution is particularly recommended for material containing chlorophyll, since 

 this will be extracted by the strong alcohol. Nuclei and leucoplasts are stained 

 a steel blue by the nigrosin. 



Potassium Alcohol. Used for bleaching sections. It may be prepared by 

 mixing a concentrated aqueous solution of potassium hydrate with 90 per cent, 

 alcohol until a sediment is formed. This is allowed to stand for 24 hours with fre- 

 quent violent shaking, and then the clear liquid is poured off and is diluted for use 

 with 2 or 3 parts of water. 



Potassium Hydrate. For general use, dissolve 5 Gm. of potassium hydrate in 

 95 mils of distilled water. This is used as a clearing agent for sections and small 

 organisms. The process of clearing may require from several hours to several days. 

 After clearing, the potash should be washed out in plenty of water, and then the 

 preparation may be neutralized with acetic acid. This will tend to make the objects 

 more opaque, and if too much is added, the objects may be cleared again by caustic 

 potash or ammonia. A dilute solution of caustic potash, as above, may be used 

 for the maceration of cork, while delicate tissues in general may be macerated by 

 boiling for a few minutes in a 50 per cent, solution of potassium hydrate in water; 

 the tissues should then be washed in water and teased out on a slide in a drop 

 of water. 



Ruthenium Red. An aqueous solution is an excellent stain for pectic substances 

 and for gums and slimes which have been derived from these. Ruthenium red 

 is not soluble in alcohol, clove oil, or glycerine, and, therefore, preparations 

 stained by it may be dehydrated and mounted in glycerine or balsam. 



Safranin. A saturated solution of safranin in alcohol should be made and this 

 should be diluted with an equal bulk of water, or with an equal bulk of a saturated 

 aqueous solution of safranin. This is an excellent general stain, and gives good 

 differentiating effects when used singly. It is one of the few stains which are par- 

 ticularly adapted to the staining of pectic compounds. It also gives beautiful 

 results in staining the cell-contents of spirogyra and other algae. The algae, after 

 fixing in a fixative containing chromic acid, should lie in the alcoholic solution 

 diluted with an equal bulk of water for 12 or 24 hours. They should be transferred 

 to 50 per cent, alcohol, to which strong alcohol is then added, drop by drop. The 

 color will begin to be extracted in the alcohol, and when the right intensity has been 

 reached, the material should be transferred to dilute glycerine, where it is to remain 

 while the glycerine slowly concentrates in a place protected against dust. Then 

 permanent mounts may be made in glycerine or glycerine-jelly. The stain given 

 by safranin is quite permanent. See also under Double Staining, and the directions 

 there given for the three-color method. 



Salicylate of Soda. A clearing reagent which for small objects is not inferior 

 to chloral hydrate is furnished by dissolving crystals of salicylate of soda in an equal 



