Virus Enrichment for Single Virus Infection by Using 3D Insulator Based Dielectrophoresis

PLOS ONE, Dec 2019

We developed an active virus filter (AVF) that enables virus enrichment for single virus infection, by using insulator-based dielectrophoresis (iDEP). A 3D-constricted flow channel design enabled the production of an iDEP force in the microfluidic chip. iDEP using a chip with multiple active virus filters (AVFs) was more accurate and faster than using a chip with a single AVF, and improved the efficiency of virus trapping. We utilized maskless photolithography to achieve the precise 3D gray-scale exposure required for fabrication of constricted flow channel. Influenza virus (A PR/8) was enriched by a negative DEP force when sinusoidal wave was applied to the electrodes within an amplitude range of 20 Vp-p and a frequency of 10 MHz. AVF-mediated virus enrichment can be repeated simply by turning the current ON or OFF. Furthermore, the negative AVF can inhibit virus adhesion onto the glass substrate. We then trapped and transported one of the enriched viruses by using optical tweezers. This microfluidic chip facilitated the effective transport of a single virus from AVFs towards the cell-containing chamber without crossing an electrode. We successfully transported the virus to the cell chamber (v = 10 µm/s) and brought it infected with a selected single H292 cell.

A PDF file should load here. If you do not see its contents the file may be temporarily unavailable at the journal website or you do not have a PDF plug-in installed and enabled in your browser.

Alternatively, you can download the file locally and open with any standalone PDF reader:

https://journals.plos.org/plosone/article/file?id=10.1371/journal.pone.0094083&type=printable

Virus Enrichment for Single Virus Infection by Using 3D Insulator Based Dielectrophoresis

Citation: Masuda T, Maruyama H, Honda A, Arai F ( Virus Enrichment for Single Virus Infection by Using 3D Insulator Based Dielectrophoresis Taisuke Masuda 0 Hisataka Maruyama 0 Ayae Honda 0 Fumihito Arai 0 Suryaprakash Sambhara, Centers for Disease Control and Prevention, United States of America 0 1 Department of Micro-Nano Systems Engineering, Graduate School of Engineering, Nagoya University, Chikusa-ku, Nagoya, Japan, 2 Department of Frontier Bioscience, Hosei University , Koganei, Tokyo , Japan We developed an active virus filter (AVF) that enables virus enrichment for single virus infection, by using insulator-based dielectrophoresis (iDEP). A 3D-constricted flow channel design enabled the production of an iDEP force in the microfluidic chip. iDEP using a chip with multiple active virus filters (AVFs) was more accurate and faster than using a chip with a single AVF, and improved the efficiency of virus trapping. We utilized maskless photolithography to achieve the precise 3D grayscale exposure required for fabrication of constricted flow channel. Influenza virus (A PR/8) was enriched by a negative DEP force when sinusoidal wave was applied to the electrodes within an amplitude range of 20 Vp-p and a frequency of 10 MHz. AVF-mediated virus enrichment can be repeated simply by turning the current ON or OFF. Furthermore, the negative AVF can inhibit virus adhesion onto the glass substrate. We then trapped and transported one of the enriched viruses by using optical tweezers. This microfluidic chip facilitated the effective transport of a single virus from AVFs towards the cellcontaining chamber without crossing an electrode. We successfully transported the virus to the cell chamber (v = 10 mm/s) and brought it infected with a selected single H292 cell. - Funding: This work was partially supported by Core Research for Evolutional Science and Technology (CREST) of Japan Science and Technology Corporation (JST). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. No additional funding was received for this study. Competing Interests: The authors have declared that no competing interests exist. Conventional analysis of viral functions is performed by using virus-infected cultured cells and this method has been considered as the most precise technique of analysis. This method provides information that is an average result of data generated from all of the cells in the population. However, the physiological state and cell cycle stage of each infected cell is different [1]. In order to quantitatively analyze viral effects, analysis of specific cell that is infected by a single virus is required. To satisfy this requirement, we previously constructed a system for the manipulation of a single virus using optical tweezers [2,3]. This single-virus infection system enabled the effective transport of a single virus from the periphery towards the cell-containing chamber. Recently this system was used to characterize the difference in influenza virus susceptibility between G1- and S/G2/M-phase cells [4]. However, bionanoparticles such as the influenza virus (shape: sphere, diameter: approximately 100 nm) tend to be present in samples at a low concentration, and a low virus number provides a limitation to the method. We therefore fabricated an active virus filter (AVF) that could enrich for viruses and modulate virus distribution, by using a dielectrophoretic force. A dielectrophoretic force is a force that is exerted on a polarizable particle under conditions of a non-uniform electric field [5,6,7]. Dielectrophoretic manipulation and accumulation of micro- and nanoparticles as well as its theoretical background were first advocated by Pohl [8]. Biological particles such as cells, bacteria, macromolecules, DNA and viruses have been extensively studied using this method [9]. There is now considerable effort being directed toward applying dielectrophoresis (DEP) for biomedical and biotechnological applications [10,11,12]. DEP has been used to trap and analyze individual cells, immobilize cells in an array format, separate different cell types (e.g. viable from dead cells), detect bacteria and manipulate viruses. Conventional electrode-based systems generate electric field gradients by applying an AC signal across two or more metallic electrodes. These systems typically use a coplanar electrode [13,14] or an interdigitated castellated microelectrode [15,16,17], and trap particles at or near the electrode surfaces. Electrode-based DEP systems have been used in the analysis of various particles for the purpose of concentration of the particles in the samples, and they exhibit high selectivity [18,19]. In insulator-based dielectrophoresis (iDEP), remote electrodes apply an electric field within a fluidic volume while insulating structures are used to distort the electric field thereby producing spatial non-uniformities [20,21,22]. They can be (...truncated)


This is a preview of a remote PDF: https://journals.plos.org/plosone/article/file?id=10.1371/journal.pone.0094083&type=printable

Taisuke Masuda, Hisataka Maruyama, Ayae Honda, Fumihito Arai. Virus Enrichment for Single Virus Infection by Using 3D Insulator Based Dielectrophoresis, PLOS ONE, 2014, 6, DOI: 10.1371/journal.pone.0094083