A METHOD FOR IN SITU CYTOPHOTOMETRIC ESTIMATION OF ABSOLUTE AMOUNT OF RIBONUCLEIC ACID USING AZURE B

Abstract

A colorimetric method is proposed for in situ cytophotometric estimation of ribonucleic acid (RNA) in biologic material, using a gelatin-RNA model system as a reference. Both specificity and stoichiometry of azure B dye binding to RNA in this model system were demonstrated. As the amount of dye binding is stoichiometric with the concentration of RNA, the regression equation generated from cytophotometric data of relative concentration ( E/d) of azure B-RNA complex as a function of concentration of RNA (picograms per cubic micron) can be used to calculate the absolute concentration (picograms per cubic micron) of RNA in biologic material. Comparison of such an estimate for nucleolar RNA in Purkinje cell nuclei of the mouse with an estimate derived from available experimental data in the literature is made to assess the validity of the reference. These estimates are in agreement, supporting the applicability of the proposed method.