Subnitaneous Tissue Stainins: 109 



'i-i 



Orcein (romeis, 1948) 

 fixation: any general fixative. 



SOLUTION : 



orcein 1 .9 gm. 



70% ethyl alcohol 100.0 ml. 



hydrochloric acid (assay 37-38%) 1.0 ml. 



procedure: 



1. Deparaffinize and hydrate slides to water; remove HgCU. 



2. Stain in orcein: 30-60 minutes. 



3. Wash briefly in distilled water. 



4. Dehydrate in 95% alcohol: 2 minutes. 



5. Differentiate in absolute alcohol until backgroimd is almost color- 

 less and elastin fibers are isolated. 



6. Rinse in fresh absolute alcohol; clear and moinit. 



results: 



elastin— red 



comments: 



Some disagreement appears in the literature concerning the solution 

 pH at which orcein is most effective. Weiss (1954) writes that orcein 

 stains only from an acid alcoholic solution between /;H 3 and 8, that 

 this puts orcein in a category between basic and acidic dyes, which 

 generally operate at the extremes of alkaline and basic pH. He there- 

 fore considers there is a formation of hydrogen bond between orcein 

 and elastin. Since this reaction takes place in acid alcohol, it is prob- 

 ably dtie to a imiquely low positive charge of elastin in such solutions 

 The use of alcoholic solutions is necessary to stabilize the positively 

 charged orcein fractions. 



Darroiv (19^2), experimenting with orcein, foimd that pH 1-2.4 

 is best for elastin staining, that above pH 2.6 collagen stains as well. 

 A dye content of 0.4% is adequate for elastin staining; a higher con- 

 centration adds to collagen staining. For a specific elastin reaction 

 when collagen is present, therefore, it is advisable to check pH of acid 

 alcohol used for the solution and to reduce the dye content to 0.4 gm. 

 per 100 ml. 



Subcutaneous Tissue Stainins: 



Subcutaneous tissue, areolar tissue, and omentum (membranes) can 

 be easily fixed for staining. A simple way is to spread a piece \vith dis- 



