326 CYTCXTHEMICAL TESTS ON EMBRYOS 



NUCLEIC ACIDS:- 



A. Toluidine Blue: 



1. Slain sections (ovary) for 20 minutes in saturated aqueous solution of Toluidine 

 Blue. 



2. Differentiate twice for 10 minutes in 95% alcohol. 



3. Mount as usual. 



Toluidine blue is taken by both nucleic acids, an orthochromatic (blue) color is 

 given only by nucleic acids, found in both the cytoplasm and nucleus. 



B. The Plasmal Reaction: 



The following procedure has been used successfully with the annphibian ovary, par- 

 ticularly the immature and post-ovulation ovary. It is based on Voss (1922 and 1931) 

 and Lison (1936). The reaction is given by a special type of phospholipid in which an 

 aldehyde group appears upon treatment with the sublimate. The procedure follows: 



1. Fix the ovary in saturated corrosive sublimate - 5 minutes. 



2. Wash in distilled water. 



3. Dehydrate, clear, embed, section (10 (jl), and then hydrate. 



4. Place directly in Feulgen reagent for 15 minutes. 



5. Wash with 3 changes of distilled water saturated with SOt. 



6. Rinse in distilled water. 



7. Mount in pure glycerine and observe under the microscope. 



Plasmalogen is a component of the cytoplasnn which gives a positive Feulgen test. 

 Being a phospholipid, the control for this test consists of fixing the ovary with Car- 

 noy's fluid and washing twice with alcohol for 15 minutes each. This extracts most 

 of the lipids. 



The Nucleal Reaction: 



This reaction depends upon the combination of the N-sulphinic acid in fuchsin-sul- 

 phurous acid with the aldehyde component released from a molecule by mild and 

 partial hydrolysis. However, it inust be remembered that the failure of any tested 

 tissue to give this reaction may be due to any of four following causes (Gardiner, 

 1935). 



1. The substance may not contain the aldehyde coniponent. 



2. The hydrolysis inay be insufficient to release the aldehyde group from the mole- 

 cule. 



3. The hydrolysis may result not only in splitting off of the aldehyde group but also 

 in its disintegration, so that it cannot react with the sulphinic acid to form the 

 new compound. 



4. The aldehyde-containing substance, and consequently the aldehyde set free on 

 hydrolysis, may be so small in quantity that, although the reaction actually 

 occurs, the compound is too minute in amount to be visible even with high mag- 

 nification. 



Gardiner (1935) says further: "Typically chromatin gives the reaction, but it does 

 not invariably do so, nor is it the only cellular substance capable of it. " Stowell 

 (1946) says: "The preponderance of evidence indicates that with the proper precau- 

 tions the Feulgen technic for thymonucleic acid is one of the most specific histo- 

 chemical reactions. " 



* The cytoplosm of cells contains principally ribonucleic acid (also known as yeast or phytonucleic acid) while the nucleus con- 

 tains largely desoxyribonucleic acid (also known as thymonucleic acid). The ribonucleic acid is transformed into thymonucleic 

 acid during development. 



