Standardization of the Feulgen reaction: the influence of chromatin condensation on the kinetics of acid hydrolysis.

The purpose of the present study was to investigate the influence of chromatin compactness on the kinetics of acid hydrolysis in the Feulgen reaction in cytology. Tissue imprints of rabbit liver, of human bronchial carcinoma and of human blood smears, fixed with alcohol, formaldehyde or with Böhm's solution with and without prior air drying, were stained with a standardized pararosanilin-Feulgen reagent. The time for hydrolysis varied between 7.5 and 120 min. The integrated optical density (IOD) of the cell nuclei was measured with an image analyzer (IBAS 2000). Cells with condensed chromatin (lymphocytes, small cell carcinoma, formaldehyde fixed cells) showed a slow increase of staining intensity and late plateau phase as compared with cells with decondensed chromatin. DNA in condensed nuclei was less susceptible to acid hydrolysis. The degree of chromatin compactness which determines the sensitivity of DNA to hydrolysis is influenced by the type of fixation, cell type and by the functional status of the cell. The conclusion is that Feulgen staining intensities of cells with different degrees of chromatin compactness cannot be compared unless measured in the respective plateau phases of the relevant hydrolysis curves which must be determined individually for each cell type.