title: International network for comparison of HIV neutralization assays: the NeutNet report.
abstract: BACKGROUND
  Neutralizing antibody assessments play a central role in human immunodeficiency virus type-1 (HIV-1) vaccine development but it is unclear which assay, or combination of assays, will provide reliable measures of correlates of protection. To address this, an international collaboration (NeutNet) involving 18 independent participants was organized to compare different assays.
  
  
  METHODS
  Each laboratory evaluated four neutralizing reagents (TriMab, 447-52D, 4E10, sCD4) at a given range of concentrations against a panel of 11 viruses representing a wide range of genetic subtypes and phenotypes. A total of 16 different assays were compared. The assays utilized either uncloned virus produced in peripheral blood mononuclear cells (PBMCs) (virus infectivity assays, VI assays), or their Env-pseudotyped (gp160) derivatives produced in 293T cells (PSV assays) from molecular clones or uncloned virus. Target cells included PBMC and genetically-engineered cell lines in either a single- or multiple-cycle infection format. Infection was quantified by using a range of assay read-outs that included extracellular or intracellular p24 antigen detection, RNA quantification and luciferase and beta-galactosidase reporter gene expression.
  
  
  RESULTS
  PSV assays were generally more sensitive than VI assays, but there were important differences according to the virus and inhibitor used. For example, for TriMab, the mean IC50 was always lower in PSV than in VI assays. However, with 4E10 or sCD4 some viruses were neutralized with a lower IC50 in VI assays than in the PSV assays. Inter-laboratory concordance was slightly better for PSV than for VI assays with some viruses, but for other viruses agreement between laboratories was limited and depended on both the virus and the neutralizing reagent.
  
  
  CONCLUSIONS
  The NeutNet project demonstrated clear differences in assay sensitivity that were dependent on both the neutralizing reagent and the virus. No single assay was capable of detecting the entire spectrum of neutralizing activities. Since it is not known which in vitro assay correlates with in vivo protection, a range of neutralization assays is recommended for vaccine evaluation.
authors.affiliation: Department of Microbiology, Dermatology and Infection, University of Lund, Lund, Sweden.
authors.forename: Eva Maria
authors.initials: EM
authors.lastname: Fenyö
authors.forename: Alan
authors.initials: A
authors.lastname: Heath
authors.forename: Stefania
authors.initials: S
authors.lastname: Dispinseri
authors.forename: Harvey
authors.initials: H
authors.lastname: Holmes
authors.forename: Paolo
authors.initials: P
authors.lastname: Lusso
authors.forename: Susan
authors.initials: S
authors.lastname: Zolla-Pazner
authors.forename: Helen
authors.initials: H
authors.lastname: Donners
authors.forename: Leo
authors.initials: L
authors.lastname: Heyndrickx
authors.forename: Jose
authors.initials: J
authors.lastname: Alcami
authors.forename: Vera
authors.initials: V
authors.lastname: Bongertz
authors.forename: Christian
authors.initials: C
authors.lastname: Jassoy
authors.forename: Mauro
authors.initials: M
authors.lastname: Malnati
authors.forename: David
authors.initials: D
authors.lastname: Montefiori
authors.forename: Christiane
authors.initials: C
authors.lastname: Moog
authors.forename: Lynn
authors.initials: L
authors.lastname: Morris
authors.forename: Saladin
authors.initials: S
authors.lastname: Osmanov
authors.forename: Victoria
authors.initials: V
authors.lastname: Polonis
authors.forename: Quentin
authors.initials: Q
authors.lastname: Sattentau
authors.forename: Hanneke
authors.initials: H
authors.lastname: Schuitemaker
authors.forename: Ruengpung
authors.initials: R
authors.lastname: Sutthent
authors.forename: Terri
authors.initials: T
authors.lastname: Wrin
authors.forename: Gabriella
authors.initials: G
authors.lastname: Scarlatti
mesh_terms: D015658:HIV Infections / Q000175:diagnosis* / Q000821:virology; D006801:Humans; D007202:Indicators and Reagents; D007391:International Cooperation; D009500:Neutralization Tests / Q000379:methods / Q000592:standards*
chemical_list: D007202:Indicators and Reagents