Control of the Ferric Ion Concentration in Iron-Aceto-Carmine Staining

The Fe⁺⁺⁺ concentration is controlled by adjusting the FeCl₃ normality of the iron-aceto-carmine staining solutions. Two stock mordant solutions are prepared by dissolving ferric chloride (FeCl₃·6H₂O; m. w. =270.31) in 45% glacial acetic acid, the normality of one being N/1, and of the other N/10. By combining aceto-carmine (preferably prepared from Merck's carmine No. 40 N. F., or from the Coleman and Bell product) and one or the other of the stock mordant solutions, a series of iron-aceto-carmine solutions is made up, each solution being of different normality for FeCl₃, depending on the proportions combined. Trial series: N/50, N/100, N/500, N/1000 and 0 N.

Tissue (spermatogenic) from one specimen is fixed in Carnoy-2, divided equally among the five iron-aceto-carmine solutions for staining, then squashed, dehydrated and mounted as usual. Subsequently the trial series may be retained or adjusted. Advantages of the method: 1) discloses quickly the optimum stain for a particular tissue type; 2) automatically gives an optimum stain to cells in different maturational stages; 3) results are reproducible in subsequent operations.

Tables and equations are provided for a number of other normalities and quantities of stain.