Separation of human metaphase chromosomes at neutral pH by velocity sedimentation at 20 times gravity

A method for the separation of large quantities of human metaphase chromosomes at neutral pH is described. The separation was performed in a specially designed sedimentation chamber which was placed in a bucket of a speed-controlled centrifuge. Flow deflectors in the chamber allowed both the undisturbed introduction of gradient and chromosomes, as well as the undisturbed fractionation of the content of the chamber. The centrifuge was run at 20 g for 1 h taking care of slow acceleration and deceleration. The slow morphological changes of the chromosomes due to ageing at neutral pH do not affect the resolving power of the sedimentation technique within this hour. This in contrast to separation of chromosomes at neutral pH by velocity sedimentation at unit gravity during 20 h, as described before. The main advantage of chromosome sorting at neutral pH is that the fractionated chromosomes are more suitable for gene transfer and gene mapping experiments.