FEULGEN NUCLEAL REACTION : I. QUANTITATIVE EXTRACTION OF FUCHSIN FROM FEULGEN-STAINED NUCLEOPROTEIN

A new extraction method for the quantitative determination of the fuchsin contained in a Feulgen-stained nucleoprotein sample has been introduced. The method is based on the following facts: (1) Treatment of a Feulgen-stained nucleoprotein sample with hot acid or alkali brings about a splitting of the linkage between fuchsin-SO₂ and the hydrolyzed nucleic acid moiety of nucleoprotein through aldehyde groups. (2) It also effectuates the formation of fuchsin from the liberated fuchsin-SO₂. (3) The fuchsin is made colorless by the treatment, but is restored to its original pink colored state when the pH of the acidic or alkaline medium is adjusted to 4.6. (4) The fuchsin, either pink colored or decolorized by alkali, can be extracted from an aqueous phase by amyl alcohol. A linear relationship was found to exist between the amount of fuchsin extracted by the FEM from a Feulgen-stained nucleoprotein sample and its DNA content. This relationship holds over a wide range of DNA concentration. From experiments utilizing this method, knowledge may be gained about the mechanism of the Feulgen reaction in situ which can lead to an improvement of the reaction in the field of cytochemistry.