Field-flow fractionation of alkali-liberated nuclear polyhedrosis virus from gypsy moth Lymantria dispar Linnaeus.
Rod-shaped viral particles of the gypsy moth nuclear polyhedrosis virus (NPV) were obtained through alkaline dissolution of inclusion bodies isolated and purified from infected hosts. The liberated viral fraction contains a wide assortment of enveloped aggregated forms as well as enveloped monomers. This complex mixture was separated by means of sedimentation field-flow fractionation (sedimentation FFF) to give a mass spectrum of the separated particles. From the elution pattern effective molecular weights could be assigned to the various resolved components. Through electron microscopy it was possible to characterize the components as monomers, dimers, etc. In parallel experiments the same viral mixture was separated by the more commonly used density gradient sedimentation technique. The two methods are compared as to time and convenience of manipulation, as well as to the amount of physicochemical information that can be extracted from each separation. The mixture of aggregate viral structures was dialyzed against a nonionic detergent solution. Sedimentation FFF of the dialysate revealed a uniform population of non-enveloped particles with no effective molecular weight which was lower than that of the initial monomer fraction. Electron microscopic observations confirmed that this lower molecular weight fraction did indeed contain rod-like structures which were thinner than the initial enveloped monomers indicating that most if not all of the enveloped material had been removed from the viral rod structures.
D000818:Animals; D002499:Centrifugation, Density Gradient; D005591:Chemical Fractionation / Q000379:methods; D007304:Insect Viruses / Q000302:isolation & purification* / Q000648:ultrastructure; D007915:Lepidoptera / Q000382:microbiology*; D008828:Microbiological Techniques; D008970:Molecular Weight; D009036:Moths / Q000382:microbiology*