A fixation method for the observation of kinetochores

Abstract

A fixation method using ethylene glycol, acetic acid and water, 1:1:1, is described. It permits light microscope studies of the kinetochore, either in unstained material using phase contrast optics, or, after post-fixation with a chromic fixative, in permanent preparations stained with crystal violet. Haemanthus, Hyacinthus, Scilla, Tradescantia and Vicia were the test materials. Literature on centromere structure is briefly reviewed. Crystal violet demonstrates compact chromatin selectively and probably stains a complex of histones with chromium. The fixative has a swelling action on the chromosomes and extracts substance from them. The kinetochore is more resistant than the chromosome body to this treatment. This resistance is due to a firmer bonding between the constituent components of the kinetochore, a firmness developed from the evolutionary adaptation of the kinetochore to resist damage by the pulling action of the spindle. The achromatic character of the kinetochore after some fixation methods may be due to the compactness of its structure which prevents penetration of dye molecules. After our fixation the kinetochore can be stained by crystal violet and the Feulgen reaction. The same compactness might also explain the differential staining of kinetochores by mitochondrial techniques.