Label-free single-particle imaging of the influenza virus by objective-type total internal reflection dark-field microscopy

Sawako Enoki; Ryota Iino; Nobuhiro Morone; Kunihiro Kaihatsu; Shouichi Sakakihara; Nobuo Kato; Hiroyuki Noji

doi:10.1371/journal.pone.0049208

Label-free single-particle imaging of the influenza virus by objective-type total internal reflection dark-field microscopy

PLoS One. 2012;7(11):e49208. doi: 10.1371/journal.pone.0049208. Epub 2012 Nov 15.

Authors

Sawako Enoki¹, Ryota Iino, Nobuhiro Morone, Kunihiro Kaihatsu, Shouichi Sakakihara, Nobuo Kato, Hiroyuki Noji

Affiliation

¹ Department of Applied Chemistry, School of Engineering, The University of Tokyo, Tokyo, Japan.

Abstract

Here we report label-free optical imaging of single particles of the influenza virus attached on a glass surface with a simple objective-type total internal reflection dark-field microscopy (TIRDFM). The capability of TIRDFM for the imaging of single viral particles was confirmed from fine correlation of the TIRDFM images with fluorescent immunostaining image and scanning electron microscopy image. The density of scattering spots in the TIRDFM images showed a good linearity against the virus concentration, giving the limit of detection as 1.2×10(4) plaque-forming units per milliliter. Our label-free optical imaging method does require neither elaborated sample preparation nor complex optical systems, offering a good platform for rapid and sensitive counting of viral particles.

Publication types

Evaluation Study
Research Support, Non-U.S. Gov't

MeSH terms

Freeze Drying / methods
Microscopy, Electron, Scanning / methods
Microscopy, Electron, Transmission / methods
Microscopy, Fluorescence / methods*
Orthomyxoviridae / ultrastructure*
Semiconductors

Grants and funding

This research was supported by the Japan Science and Technology Agency for Core Research for Evolutional Science and Technology (CREST), and supported in part by Grant-in-Aid for Scientific Research (B) (23310087 to NM) and by a Health Labor Science Research Grant (Nano-001 to NM). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.