Purification of viruses by electrophoresis in sucrose concentration gradients. 1972.

Abstract

Zone electrophoresis (ZE) in concentration gradients of sucrose proved to be a powerful technique in the purification of viruses although it is seldomly used as a single step for the separation of the infective agent from extraneous matter but usually as a final procedure. No difficulties were experienced in the purification of plant viruses with ZE possibly because of the resistance of the agents to the chloroform butanol treatment which the impure plant viruses received prior to the ZE. Likewise no difficulties were encountered during purification of human and animal picorna viruses because they also are resistant to chloroform or chloroform-butanol treatment prior to ZE. Purification of animal and possibly human reoviridae ZE as a final step is involved as they are associated with or even attached to the host cell membranes which are lipoidal in nature. Because the lipoidal material on the virus has an affinity for organic solvents the virus particles are caught in the emulsion when an extract of the crude virus material is shaken with the organic solvent. With these viruses polyethylene glycol M(r) 6000 (PEG) is often helpful as a means of displacing most of the host proteins as a step in the partial purification prior to ZE. Neurotropic African horse sickness virus which presented many problems to overcome was rendered free of extraneous proteins to the extent that electron micrographs could be made of it. Neurotropic Rift Valley fever virus was similarly difficult to purify as it was always associated with mouse brain components. This virus was finally purified from infected mouse blood in which the virus particles were not associated with host cell debris and other extraneous proteins. Zone electrophoresis experiments were not conducted on viruses with large particle sizes e.g. myxoviruses because they could be purified without a great deal of effort using polymer displacement, and thin layer centrifugation. Zone electrophoresis as a means of final purification was vividly displayed with the purification of five viruses of the larvae of the pine emperor moth, Nudaurelia cytheria cytheria.