CONTROLLED CHROMATION

Abstract

Controlled chromation represents another step in the development of dichromate oxidation as a cytochemical technique, to which previous contributions have been made by Weigert, Smith, Dietrich, Ciaccio and Baker. Recognition of the critical importance of controlling pH, in addition to the regulation of temperature, time, and concentration, allows voluntary adjustment of these variables to fit each particular problem. Control of chromation is equally important whether it is exercised on fresh tissues, frozen sections, or tissues previously subjected to the action of fixatives or solvents. By using graded intensities of chromation, differential visualization of a series of cell constituents may be achieved. Judicious combination of controlled chromation with the use of other reagents, including solvents, offers the possibility of developing analytical methods for the study of a great variety of lipid and non-lipid tissues components. The relative ease with which phospholipids, such as those of the Golgi apparatus, are chromated allows their routine visualization to be accomplished after chromation at pH 3.5 for 18 hours at 56°C. with 2.5% potassium dichromate, followed by paraffin embedding. Alternate sections stained with Sudan black B and with hematoxylin can be ready for study within twenty four hours of the collection of the tissue. Chromated lipids are stained by both Sudan black B and by hematoxylin, non-chromated lipids only by Sudan black B, and non-lipid chromated substances only by hematoxylin. Other dyes may be used in place of Sudan black B and hematoxylin when advantageous. The staining of chromated tissue with hematoxylin can be controlled by the addition of any one of several reagents to the staining solution, so as to restrain the formation of hematoxylin lakes by minor concentrations of chromium. Differentiation is consequently accomplished during staining and the entire process can be completed on paraffin sections at 56°C. in two hours.