Tests for Nucleic Acid Selectivity of Einarson's Gallocyanin-Chrome Alum Stain on Monolayers of Strain L-929 Mouse Fibroblasts

Abstract

L-929 fibroblasts, fixed on coverslips, were stained with gallocyanin-chrome alum after various treatments for removal of nucleic acid or for methylation or deamination. For nucleic acid, trichloroacetic acid and NaCl extractions and sequential incubation in DNase and RNase yielded cells unstainable with the dye complex. Methylated cells showed no cytoplasmic staining and a reduced nuclear staining, compared with the unblocked controls. Deamination had little effect. All results were dependent on the types of fixative used, times and temperatures of incubation, and in the case of nucleases, their concentration. Conventional dehydration and melted paraffin infiltration was associated with little or no staining of deaminated cells and intense staining of methylated cells. The paraffin effects were also dependent on fixatives. The evidence shows that gallocyanin-chrome alum combines with groups (presumably phosphate or carboxyl, or both) which are blocked by methylation, and which can be removed from L cells by sequential RNase and DNase treatment.