Characterization of vesicular stomatitis virus populations by tunable resistive pulse sensing

Abstract

Although transmission electron microscopy (TEM) has historically been the method of choice to estimate concentrations of virus and virus-like particles, these measures can often be time-consuming and labor-intensive to perform. Tunable resistive pulse sensing (TRPS) is an emerging method that applies principles of Coulter counting to nanoscale particles and may provide a simpler and higher-throughput alternative to TEM for the quantitation of virus populations. To assess the performance of TRPS compared to TEM, the samples of polymer spheres at a diameter of 100 nm and vesicular stomatitis virus (VSV) were characterized using both techniques. TRPS was able to quantify concentrations down to 107 particles per ml, providing nearly 50-fold larger measurement range, and more reproducible counts than TEM. Total-to-infectious particle ratio of VSV populations as measured by TRPS and plaque assay suggested that each VSV particle is infectious. In addition to particle counts, TRPS successfully measured particle size distributions based on hundreds of particles. Such high throughput sustained by TRPS can assist quantitative characterization of virus populations.

Inhibition of infection spread by co-transmitted defective interfering particles

“…The DIP-GFP working stock was prepared by infecting BHK cells transfected with pMD2.G. The particle concentrations of the DIP and DIP-GFP stocks were determined via tunable resistive pulse sensing [ 60 , 61 ]. The concentration determined by these particle counts were similar to those determined by qPCR [ 56 ].…”

“…At 24 hpi the cells were imaged on a Nikon TE Eclipse 300 inverted microscope, and the supernatant was collected and stored at -80°C. The number of infectious virus particles in the supernatant was determined by plaque assay, and the total particle counts, which includes infectious and non-infectious particles, were determined using tunable resistive pulse sensing via a qViro (IZON Science, Christchurch, NZ) [ 60 ].…”

Quantification of Virus Particles Using Nanopore-Based Resistive-Pulse Sensing Techniques

“…Farkas and coworkers counted rotavirus (75 nm) using both qRT-PCR and qNano to evaluate the purification of the viruses by size-exclusion chromatography (Farkas et al, 2013 ). Akpinar and Yin counted vesicular stomatitis virus (70 × 200 nm) using both TRPS and a plaque assay (Akpinar and Yin, 2015 ). The average total to infectious particle ratio was calculated as 2.91 ± 1.42.…”

Tunable Resistive Pulse Sensing: Potential Applications in Nanomedicine

“…They were able to measure the size and density of human immunodeficiency virus and Epstein-Barr virus. Using vesicular stomatitis virus (VSV) and 100nm polymer spheres as size controls, Akpinar & Yin [75] has shown that TRPS was able to quantify concentrations down to 107 particles per ml which was 50 fold lower than the detection limit of TEM. TRPS produced more reproducible counts than TEM.…”