Factors affecting the efficiency of immunogold labelling of plant virus antigens in thin sections

Sections of pellets of six purified plant viruses with three different morphologies were used to examine different technical aspects of the immunogold labelling (IGL) technique. The results showed that fixation by glutaraldehyde alone was better than with osmium tetroxide post-fixation, and that Decon 75 was the best of the pretreatments tried. The study showed that different virus homologous antisera gave different results in IGL tests, and that longer incubation times with both antiserum and gold probe gave higher label densities without any increase in background label. Also, cross-absorption of the virus antisera with healthy host protein before use gave cleaner backgrounds and thus higher specificity. The work also examined the relationship between label density and amounts of visible virus. There was no correlation between the numbers of virus particles seen in sections and the numbers of gold particles; moreover, there was no apparent relationship between label density and the orientation or distribution of the virus particles in the section. The role of the embedding resin and its polymerisation temperature are also discussed.