Use of protein A in the serum-in-agar diffusion method in immune electron microscopy for detection of virus particles in cell culture.
A modified technique using protein A in the serum-in-agar (SIA) method for immune electron microscopy (IEM) was presented. Grids coated with staphylococcal protein A were floated on samples mounted on agar containing 2% antiserum and incubated at 37 C, for 60 min. After washing and staining, the grids were observed in an electron microscope. The effects of protein A on virus detection were evaluated using poliovirus and bovine rotavirus infected cell culture fluids. The results showed that the technique using protein A (PA-SIA) had at least 10-fold higher sensitivity for virus detection than the original SIA. The optimal concentration of protein A was 1 to 10 micrograms/ml for coating the grids to trap virus particles. The PA-SIA method was also compared with immunosorbent electron microscopy (ISEM). The former showed higher or at least the same sensitivity and some advantages in detecting antigen-antibody reaction than the latter method. These results indicate that our PA-SIA method may be superior to other IEM techniques presented previously for the detection and identification of viruses.
D000327:Adsorption; D000914:Antibodies, Viral / Q000276:immunology; D000936:Antigen-Antibody Complex; D005779:Immunodiffusion; D008854:Microscopy, Electron / Q000379:methods; D013205:Staphylococcal Protein A; D014780:Viruses / Q000032:analysis* / Q000276:immunology / Q000648:ultrastructure
D000914:Antibodies, Viral; D000936:Antigen-Antibody Complex; D013205:Staphylococcal Protein A