Journal
JOURNAL OF VIROLOGICAL METHODS
Volume 163, Issue 2, Pages 336-343Publisher
ELSEVIER SCIENCE BV
DOI: 10.1016/j.jviromet.2009.10.020
Keywords
Virus entry; Influenza virus; Ebola virus; Marburg virus; Flow cytometry; Microscopy
Funding
- NIH-NCI [5R24 CA09582304]
- NSF [DBI-9724504]
- NIH [1 S10 RR0 9145-01]
- Center for Research on Influenza Pathogenesis (CRIP)
- National Institute for Allergy and Infectious Diseases (NIAID) [HHSN266200700010C, U01 A170469, P01 A158113, 1F32A1081428]
- Northeast Biodefense Center
- Center for Investigating Viral Immunity and Antagonism (CIVIA) [U54AI57158, U19AI62623]
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A viral entry assay where a beta-lactamase reporter protein fused to the influenza matrix protein-1 (BlaM1) is packaged as a structural component into influenza virus-like particles (VLPs) is described The. Bla reporter is released upon fusion with target cells and can be detected in live cells by flow cytometry, microscopy, or fluorometric plate reader for utility in high-throughput screening approaches. The production of BlaM1 VLPs and subsequent transfer of Bla activity to target cells requires the presence of influenza hemagglutinin (HA) and neuraminidase (NA). In addition, transfer of Bla by the VLPs can be blocked by an influenza neutralizing antibody, is impeded by a chemical inhibitor of influenza virus entry, and requires HA that is cleaved by a protease specific for its cleavage site. An analogous VLP system also was developed for Ebola (EBOV) and Marburg (MARV) viruses, demonstrating that this straightforward assay has broad application for studying the entry steps of enveloped viruses, especially those that require high levels of biosafety containment. (C) 2009 Elsevier B.V. All rights reserved.
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