title: Capturing enveloped viruses on affinity grids for downstream cryo-electron microscopy applications.
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.
authors.affiliation: Department of Pediatrics, School of Medicine, Division of Pediatric Infectious Diseases, Emory University, Children's Healthcare of Atlanta, Atlanta, GA 30322, USA.
authors.forename: Gabriella
authors.initials: G
authors.lastname: Kiss
authors.affiliation: Department of Pediatrics, School of Medicine, Division of Pediatric Infectious Diseases, Emory University, Children's Healthcare of Atlanta, Atlanta, GA 30322, USA.
authors.forename: Xuemin
authors.initials: X
authors.lastname: Chen
authors.affiliation: Center for Inflammation, Immunity & Infection, Georgia State University, Atlanta, GA 30303, USA.
authors.forename: Melinda A
authors.initials: MA
authors.lastname: Brindley
authors.affiliation: Department of Microbiology and Immunology, School of Medicine, Emory University, Atlanta, GA 30322, USA.
authors.forename: Patricia
authors.initials: P
authors.lastname: Campbell
authors.affiliation: USDA, ARS, Southeast Poultry Research Laboratory, Athens, GA 30605, USA.
authors.forename: Claudio L
authors.initials: CL
authors.lastname: Afonso
authors.affiliation: School of Biology, Georgia Institute of Technology, Atlanta, GA 30332, USA.
authors.forename: Zunlong
authors.initials: Z
authors.lastname: Ke
authors.affiliation: Department of Pediatrics, School of Medicine, Division of Pediatric Infectious Diseases, Emory University, Children's Healthcare of Atlanta, Atlanta, GA 30322, USA.
authors.forename: Jens M
authors.initials: JM
authors.lastname: Holl
authors.affiliation: Department of Pediatrics, School of Medicine, Division of Pediatric Infectious Diseases, Emory University, Children's Healthcare of Atlanta, Atlanta, GA 30322, USA.
authors.forename: Ricardo C
authors.initials: RC
authors.lastname: Guerrero-Ferreira
authors.affiliation: Department of Microbiology and Immunology, School of Medicine, Emory University, Atlanta, GA 30322, USA.
authors.forename: Lauren A
authors.initials: LA
authors.lastname: Byrd-Leotis
authors.affiliation: Department of Microbiology and Immunology, School of Medicine, Emory University, Atlanta, GA 30322, USA.
authors.forename: John
authors.initials: J
authors.lastname: Steel
authors.affiliation: Department of Microbiology and Immunology, School of Medicine, Emory University, Atlanta, GA 30322, USA.
authors.forename: David A
authors.initials: DA
authors.lastname: Steinhauer
authors.affiliation: Department of Pediatrics, School of Medicine, Division of Pediatric Infectious Diseases, Emory University, Children's Healthcare of Atlanta, Atlanta, GA 30322, USA.
authors.forename: Richard K
authors.initials: RK
authors.lastname: Plemper
authors.affiliation: Virginia Tech Carilion Research Institute, Roanoke, VA 24016, USA.
authors.forename: Deborah F
authors.initials: DF
authors.lastname: Kelly
authors.affiliation: Department of Pediatrics, School of Medicine, Division of Pediatric Infectious Diseases, Emory University, Children's Healthcare of Atlanta, Atlanta, GA 30322, USA.
authors.forename: Paul W
authors.initials: PW
authors.lastname: Spearman
authors.affiliation: Department of Pediatrics, School of Medicine, Division of Pediatric Infectious Diseases, Emory University, Children's Healthcare of Atlanta, Atlanta, GA 30322, USA.
authors.forename: Elizabeth R
authors.initials: ER
authors.lastname: Wright
mesh_terms: D020285:Cryoelectron Microscopy / Q000379:methods*; D009571:Nitrilotriacetic Acid / Q000031:analogs & derivatives; D009942:Organometallic Compounds; D014771:Virion / Q000737:chemistry / Q000302:isolation & purification*; D014780:Viruses / Q000737:chemistry / Q000302:isolation & purification*
chemical_list: D009942:Organometallic Compounds; C088321:nickel nitrilotriacetic acid; D009571:Nitrilotriacetic Acid