A study of the effect of viscosity on the resolution of bands in the preparative ultracentrifuge
Band velocity experiments in the preparative ultracentrifuge are usually performed in a linear density gradient established by mixing a 5% and a 20% sucrose solution. This stabilizing density gradient introduces a significant viscosity gradient which in turn may effect the resolving power of the sed...
| Summary: | Band velocity experiments in the preparative ultracentrifuge are usually performed in a linear density gradient established by mixing a 5% and a 20% sucrose solution. This stabilizing density gradient introduces a significant viscosity gradient which in turn may effect the resolving power of the sedimentation velocity experiment. The fast components travel on the average through more viscous solutions than do the slow components.
In this investigation the effect of viscosity gradients
was examined by experiments in which density stability was
achieved by using a cesium chloride density gradient and the viscosity was independently controlled by introducing a uniform sucrose concentration or a positive or negative sucrose gradient in the cesium chloride gradient.
Experiments performed with a mixture of tritiated thymidine labeled polyoma DNA components I, II, and III showed
that the separation of band maxima increased as the direction of the viscosity gradient was changed from positive to zero to negative with respect to field. On the other hand the band widths increased in the same order. The resolution in terms of the separation between the bands divided by the sum of the band widths was clearly lowest with the negative viscosity gradient and was approximately the same for the positive and zero viscosity gradients. |
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