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Capturing Enveloped Viruses on Affinity Grids for Downstream Cryo-Electron Microscopy Applications

Published online by Cambridge University Press:  26 November 2013

Gabriella Kiss ,
Xuemin Chen ,
Melinda A. Brindley ,
Patricia Campbell ,
Claudio L. Afonso ,
Zunlong Ke ,
Jens M. Holl ,
Ricardo C. Guerrero-Ferreira ,
Lauren A. Byrd-Leotis  and
John Steel
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Gabriella Kiss
Affiliation:
Department of Pediatrics, School of Medicine, Division of Pediatric Infectious Diseases, Emory University, Children's Healthcare of Atlanta, Atlanta, GA 30322, USA
Xuemin Chen
Affiliation:
Department of Pediatrics, School of Medicine, Division of Pediatric Infectious Diseases, Emory University, Children's Healthcare of Atlanta, Atlanta, GA 30322, USA
Melinda A. Brindley
Affiliation:
Center for Inflammation, Immunity & Infection, Georgia State University, Atlanta, GA 30303, USA
Patricia Campbell
Affiliation:
Department of Microbiology and Immunology, School of Medicine, Emory University, Atlanta, GA 30322, USA
Claudio L. Afonso
Affiliation:
USDA, ARS, Southeast Poultry Research Laboratory, Athens, GA 30605, USA
Zunlong Ke
Affiliation:
School of Biology, Georgia Institute of Technology, Atlanta, GA 30332, USA
Jens M. Holl
Affiliation:
Department of Pediatrics, School of Medicine, Division of Pediatric Infectious Diseases, Emory University, Children's Healthcare of Atlanta, Atlanta, GA 30322, USA
Ricardo C. Guerrero-Ferreira
Affiliation:
Department of Pediatrics, School of Medicine, Division of Pediatric Infectious Diseases, Emory University, Children's Healthcare of Atlanta, Atlanta, GA 30322, USA
Lauren A. Byrd-Leotis
Affiliation:
Department of Microbiology and Immunology, School of Medicine, Emory University, Atlanta, GA 30322, USA
John Steel
Affiliation:
Department of Microbiology and Immunology, School of Medicine, Emory University, Atlanta, GA 30322, USA
David A. Steinhauer
Affiliation:
Department of Microbiology and Immunology, School of Medicine, Emory University, Atlanta, GA 30322, USA
Richard K. Plemper
Affiliation:
Department of Pediatrics, School of Medicine, Division of Pediatric Infectious Diseases, Emory University, Children's Healthcare of Atlanta, Atlanta, GA 30322, USA Center for Inflammation, Immunity & Infection, Georgia State University, Atlanta, GA 30303, USA
Deborah F. Kelly
Affiliation:
Virginia Tech Carilion Research Institute, Roanoke, VA 24016, USA
Paul W. Spearman
Affiliation:
Department of Pediatrics, School of Medicine, Division of Pediatric Infectious Diseases, Emory University, Children's Healthcare of Atlanta, Atlanta, GA 30322, USA
Elizabeth R. Wright*
Affiliation:
Department of Pediatrics, School of Medicine, Division of Pediatric Infectious Diseases, Emory University, Children's Healthcare of Atlanta, Atlanta, GA 30322, USA
*
*Corresponding author. E-mail: [email protected]
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Abstract

Electron microscopy (EM), cryo-electron microscopy (cryo-EM), and cryo-electron tomography (cryo-ET) are essential techniques used for characterizing basic virus morphology and determining the three-dimensional structure of viruses. Enveloped viruses, which contain an outer lipoprotein coat, constitute the largest group of pathogenic viruses to humans. The purification of enveloped viruses from cell culture presents certain challenges. Specifically, the inclusion of host-membrane-derived vesicles, the complete destruction of the viruses, and the disruption of the internal architecture of individual virus particles. Here, we present a strategy for capturing enveloped viruses on affinity grids (AG) for use in both conventional EM and cryo-EM/ET applications. We examined the utility of AG for the selective capture of human immunodeficiency virus virus-like particles, influenza A, and measles virus. We applied nickel-nitrilotriacetic acid lipid layers in combination with molecular adaptors to selectively adhere the viruses to the AG surface. This further development of the AG method may prove essential for the gentle and selective purification of enveloped viruses directly onto EM grids for ultrastructural analyses.

Keywords

cryo-electron microscopy (cryo-EM)cryo-electron tomography (cryo-ET)affinity grid (AG)enveloped virus
Type
Techniques, Software, and Instrumentation Development
Information
Microscopy and Microanalysis , Volume 20 , Issue 1 , February 2014 , pp. 164 - 174
Copyright
Copyright © Microscopy Society of America 2014 

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