AN EVALUATION OF THE BARRNETT AND SELIGMAN PROCEDURE WHEN USED TO DETERMINE CHANGES IN THE RATIO OF NUCLEAR PROTEIN-BOUND DISULFIDE TO SULFHYDRYL GROUPS DURING MITOSIS

Abstract

Using a modification of the Barrnett and Seligman histochemical procedure, the proportion of protein-bound S—S/SH Was measured in the chromatin of onion root tip cells at three different stages of mitosis as well as in the nucleolus. Inconsistencies between experiments led to studies of the nature of the histochemical reaction before and after sections were reduced in thioglycolate. Different fixatives and ribonuclease were tested for their effects on the outcome of the reaction. Some deviation from the change in S—S/SH ratio during mitosis found after ethanol-acetic fixation occurs in material fixed in lO% trichloroacetic acid, 5% acetic acid and saturated HgCl₂. Ribonuclease treatment was found to increase significantly SH stain in the nucleolus, but not significantly in the resting nucleus, metaphase, or telophase chromatin. The possibility that nucleic acids may modify the SH reaction after 5% acetic acid fixation is raised. Further, the data are interpreted to indicate that saturated HgCl₂ as well as the histochemical reaction itself makes disulfide bonds stainable in the absence of reduction. After fixation in standard ethanol-acetic acid mixtures the S—S/SH ratio is highest at interphase, lowest at telophase, and intermediate at metaphase.