Localization of chromosome scaffold by transmitted- and incident-light fluorescence microscopy

In order to demonstrate the chromosome scaffold the spermatocytes of a grasshopper were fixed in acetomethanol, treated with alkaline 2XSSC and stained with AgNO3 solution. The air-dried preparations were fluorochromed with acridine orange (AO) and excited with either transmitted or incident beam of UV for fluorescence microscopic study. Under transmitted-light the scaffold is very clearly visible as non-fluorescent dark silver deposition along the middle of the chromosomes. However, under incident-illumination the scaffold is invisible and only the fluorescence of the epichromatin is observed. This indicates that the scaffold is situated longitudinally in the centre of a solid cylinder of epichromatin made of DNA-histone.