title: Improved method for counting virus and virus like particles.
abstract: An improved method for counting virus and virus like particles by electron microscopy (EM) was developed. The procedure involves the determination of the absolute concentration of pure or semi-pure particles once deposited evenly on EM grids using either centrifugation or antibody capture techniques. The counting of particles was done with a Microfiche unit which enlarged approximately 50 x the image of particles on a developed negative film which had been taken at a relatively low magnification (2500 x) by EM. Initially, latex particles of a known concentration were counted using this approach, to prove the accuracy of the technique. The latex particles were deposited evenly on an EM grid using centrifugation (Modified Beckmen EM-90 Airfuge technique). Subsequently, recombinant Bluetongue virus (BTV) core-like particles (CLPs) captured by a Monoclonal antibody using a novel sample loading method were counted by the Microfiche unit method and by a direct EM method. Comparison of the simplified counting method developed with a conventional method, showed good agreement. The method is simple, accurate, rapid, and reproducible when used with either pure particles or with particles from crude cell culture extracts.
authors.affiliation: Department of Chemical Engineering, University of Queensland, Brisbane, Australia.
authors.forename: Y Z
authors.initials: YZ
authors.lastname: Zheng
authors.forename: R
authors.initials: R
authors.lastname: Webb
authors.forename: P F
authors.initials: PF
authors.lastname: Greenfield
authors.forename: S
authors.initials: S
authors.lastname: Reid
mesh_terms: D000911:Antibodies, Monoclonal / Q000276:immunology; D015153:Blotting, Western; D001820:Bluetongue virus / Q000276:immunology / Q000302:isolation & purification / Q000648:ultrastructure; D002498:Centrifugation; D004591:Electrophoresis, Polyacrylamide Gel; D008854:Microscopy, Electron / Q000379:methods*; D016253:Microscopy, Immunoelectron / Q000379:methods*; D016624:Negative Staining; D015203:Reproducibility of Results; D013048:Specimen Handling / Q000379:methods; D014771:Virion / Q000302:isolation & purification* / Q000648:ultrastructure
chemical_list: D000911:Antibodies, Monoclonal