Plaque assay. BHK cells were plated into six-well plates and cultured to 90% confluence. Culture medium was removed from each well and replaced with 200 ll of serially diluted viral samples. The inoculated monolayers were returned to the incubator for 1 h to allow virus adsorption. The plates were rocked gently every 20 min. At the end of the adsorption period, the inoculum was removed from each monolayer sample and then replaced with 2 ml of agar overlay. The agar overlay consisted of 0.6% weight/volume (w/v) agar (Agar Nobel, Difco, BD Diagnostic Systems, http://www.bd.com). 5-Bromo-29-deoxyuridine (B5002, Sigma) was added, at 100 lg/ml, to the agar overlay of N3-and N4-infected samples to enhance plaque formation. Following agar addition, the plates were allowed to cool at room temperature for 30 min, returned to the incubator and incubated for 24 h, and then each sample was fixed with 2 ml of fixative for 3 h at room temperature. The fixative consisted of 4% (w/v) paraformaldehyde (VWR) and 5% (w/v) sucrose (Sigma) in 10 mM phosphate buffered saline (PBS, Sigma) of pH 7.4. The agar overlay was then removed, and each sample was rinsed twice with 2 ml of PBS. Gentian violet diluted in methanol (0.01% (w/v), Sigma) was used, at 1 ml each, to stain the samples. Model development. Using algebraic and differential forms of equations, our mathematical model aims to account for established molecular processing steps in the development of VSV. Most model parameters were extracted from the literature. However, five parameters were obtained by fitting our simulation results to experimental data that were from the literature and our own experiments. Key model parameters are given in Table 1 , and detailed descriptions of the model and parameter estimation process are provided below, and in Protocol S1 and Figures S2 and S3 , respectively.