Chromosome banding: specification of structural features of dyes giving rise to G-banding.

Metaphase chromosomes were stained in a routine G-banding procedure with 39 basic dyes of varied structures substituted for the Giemsa stain. Staining outcomes were categorized as: overstained, differentially stained, trivially or unstained. Certain structural features of the dyes were described numerically, namely, largest conjugated fragment (LCF), conjugated bond number (CBN) and cationic weight. The staining outcomes were compared to these numerical structural parameters, and structure--staining correlations sought. Dyes with large conjugated systems (and high LCF values) were seen to be overstained; dyes with low LCF values were often non-staining. At intermediate LCF values, the more hydrophobic dyes (with high Hansch pi values) stained differentially; the more hydrophilic dyes failed to stain. Expressed numerically, 89% of the dyes with the following characteristics stained differentially: 30 greater than or equal to LCF greater than or equal to 10; Hansch pi greater than -5.0. It was concluded that contributions to dye-chromosome affinity included coulombic forces and van der Waals attractions and that the selectivity of G-banding was largely due to hydrophobic bonding. Induction of bands could be due to the loss of hydrophilic histones, amplifying underlying variations in the hydrophobic-hydrophilic character of the chromosome structure. Relatively hydrophobic sites include AT-rich DNA and disulphide-rich proteins. The effects on Romanowsky G-banding of chemically modifying chromosomes were in keeping with this model. Overstaining resulted from formation of either hydrophobic or conjugated derivatives or both, whereas trivial or non-staining arose from the formation of hydrophilic derivatives. Intriguingly, the efficacy of the dyes used for Q-banding also correlated positively with their hydrophobic character.