Characterization of purified VSV virion preparations. (A) Electron micrographs of the purified virion preparations from BHK21 (two different fields are shown), 4T1 and A549. Virions were absorbed to carbon-formvar coated grids and negatively stained with 2% uranyl acetate. (B) Infectivity (PFU/ml) of purified virions shown in (A) was calculated by standard plaque assay on BHK21 cells. Total protein concentration of these preparations was determined using a Bradford assay, and infectivity per total protein (PFU/μg) was calculated based on these two values. (C) Total RNA was extracted from uninfected (mock) or infected BHK21 cells or from purified virion samples containing 25 μg of protein, and analyzed by Northern blotting. RNA was separated on a 1.5% agarose-formaldehyde gel, transferred to a nylon membrane and detected using a probe complimentary to the 3' end of VSV genome. (D) 50 μg of total protein from each purified virion preparation was separated on a 10% SDS-PAGE gel, and stained with Coomassie Brilliant Blue R250. Numbered boxes indicate the gel segments cut out and analyzed by mass spectrometry.