Enhanced light microscopy visualization of virus particles from Zika virus to filamentous ebolaviruses

PLoS One. 2017 Jun 26;12(6):e0179728. doi: 10.1371/journal.pone.0179728. eCollection 2017.

Abstract

Light microscopy is a powerful tool in the detection and analysis of parasites, fungi, and prokaryotes, but has been challenging to use for the detection of individual virus particles. Unlabeled virus particles are too small to be visualized using standard visible light microscopy. Characterization of virus particles is typically performed using higher resolution approaches such as electron microscopy or atomic force microscopy. These approaches require purification of virions away from their normal millieu, requiring significant levels of expertise, and can only enumerate small numbers of particles per field of view. Here, we utilize a visible light imaging approach called Single Particle Interferometric Reflectance Imaging Sensor (SP-IRIS) that allows automated counting and sizing of thousands of individual virions. Virions are captured directly from complex solutions onto a silicon chip and then detected using a reflectance interference imaging modality. We show that the use of different imaging wavelengths allows the visualization of a multitude of virus particles. Using Violet/UV illumination, the SP-IRIS technique is able to detect individual flavivirus particles (~40 nm), while green light illumination is capable of identifying and discriminating between vesicular stomatitis virus and vaccinia virus (~360 nm). Strikingly, the technology allows the clear identification of filamentous infectious ebolavirus particles and virus-like particles. The ability to differentiate and quantify unlabeled virus particles extends the usefulness of traditional light microscopy and can be embodied in a straightforward benchtop approach allowing widespread applications ranging from rapid detection in biological fluids to analysis of virus-like particles for vaccine development and production.

Publication types

  • Evaluation Study
  • Validation Study

MeSH terms

  • Animals
  • Ebolavirus / ultrastructure*
  • Equipment Design
  • Humans
  • Microscopy, Electron, Scanning
  • Microscopy, Interference / instrumentation
  • Microscopy, Interference / methods*
  • Microscopy, Ultraviolet / instrumentation
  • Microscopy, Ultraviolet / methods*
  • Vaccinia virus / ultrastructure
  • Vesiculovirus / ultrastructure
  • Virion / ultrastructure*
  • Zika Virus / ultrastructure*

Grants and funding

This work was funded in part by the National Institutes of Allergy and Infectious Diseases through awards to John H Connor (R01AI1096159) and David S Freedman (1R43AI118254). These funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. nanoView Diagnostics Inc provided support in the form of salaries for authors [GD, DS, AR, DB] and played a role in the study design, data collection and analysis, decision to publish, and prepare of the manuscript. The specific roles of these authors are articulated in the ‘author contributions’ section.