lOf) Stniyjs nnd Staining (chap. 10) 



A weak acid will not combine with nuclear elements in sufficient quan- 

 tity to produce efficient staining. Some form of mordanting (page 107) 

 is required to form a base from this dye, whi( h will then stain the acidic 

 nuclear elements. The most commonly used mordants are alum salts of 

 alumininn, potassium or iron, as already mentioned. 



Baker [19^8) recommends oxidation with sodium iodate for pre- 

 paring hematoxylin solutions. Start with a wholly unoxidized he- 

 matoxylin dye powder, not hematein powder. Solutions started with 

 the latter tend to lose their strength by flocculation (sedimentation) 

 of the products of oxidation. The hematoxylin dye when in solution 

 should be only partly oxidized by the sodium iodate. Use less chemical 

 than would be required for complete oxidation; about one fourth to 

 one half of the full amoinit of oxidizer is adequate. The solution will 

 then continue to gradually ripen by atmospheric oxygen and thereby 

 maintain its strength. Such solutions are allowed to ripen slowly, six 

 weeks or more. {Mayer's hematoxylin is an example of this type.) They 

 will produce brilliant staining for many months. 



The rate of oxidation is also affected by the solvent. A neutral 

 aqtieous solution forms hematein in a fe\v hotirs; an acid solution does 

 this more slowly and an alkaline solution more rapidly. Alcoholic solu- 

 tions are slow, and the addition of glycerine retards them even more. 

 Color changes which take place in a stock soliuion indicate its efficiency. 

 The changes, with no mordant present, are from water white through 

 lilac, bright purple, deep purple, red, orange red, orange brown to 

 brown. At the purple stage, the solution is most vigorous; at the red 

 stages less so; and at the brown stage it is no longer useful. The lifetime 

 of alcoholic solutions is five times greater than that of aqueotis ones. 

 {Cole, 1943) 



Hematoxylin is an exceedingly powerful dye with various shades of 

 staining from purples, through blues and into blue-blacks. The iron- 

 mordanted form is one of the most valuable dyes for mitotic study, and 

 gives to the chromatin a precise black or blue-black color. This black 

 color is the result of the presence in hematoxylin of some tannin, and 

 the latter in combination with iron salts produces a lasting black color. 



Other natural dyes are saffron from stigmas of Crocus; indigo from 

 plants of the genus, Indigofera; berberine from barberry; orcein and 

 LITMUS from the lichens, Lecanora and Rocella; and brazilin from 

 brazilwood — a redwood tree of the tropics. Orcein, a specific dye for 

 elastin (present in elastic fibers), is prepared by boiling the plants in 

 water. The lecanoric acid in them splits to produce orcinol — a resorcinol 



