Nomarski-optical studies of human chromosomes R-banded with barium hydroxide

The morphologic changes occurring in human chromosomes during R-banding by Ba(OH)2 treatment were followed with the aid of bright-field and Nomarski interference contrast microscopy. It was found that the hot Ba(OH)2 pretreatment alone, i.e., without staining, caused a pattern of transverse ridges in the chromosomes that clearly corresponded to positive R-band regions. No chromosomal collapse could be seen during any stage of the R-banding procedure. Thus these events contrast with those observed in G-band formation with trypsin, where complete chromosomal collapse occurs after pretreatment and where staining is necessary to induce G-band ridges. The possible mechanism of R-band induction by Ba(OH)2 is discussed. It is proposed that the R-band ridges arise as a result of chromatin loss from the interband regions during the hot alkaline pretreatment.