Home › Immunology › Modeling The Lifecycle Of Ebola Virus Under Biosafety Level 2 Conditions With Virus-like Particles Containing Tetracistronic Minigenomes
Immunology JoVE (Open Access) Citable · DOI

Modeling The Lifecycle Of Ebola Virus Under Biosafety Level 2 Conditions With Virus-like Particles Containing Tetracistronic Minigenomes

DOI: 10.3791/52381-v
What you'll learn
  • ✓ Produce tetracistronic minigenome-containing virus-like particles safely under BSL-2 conditions
  • ✓ Infect target cells with trVLPs and harvest samples for downstream analysis
  • ✓ Measure reporter activity to assess Ebola lifecycle progression without infectious virus
Protocol

Work with infectious Ebola viruses is restricted to biosafety level 4 laboratories. Tetracistronic minigenome-containing replication and transcription-competent virus like particles (trVLPs) represent a lifecycle modeling system that allows us to safely model multiple infectious cycles under biosafety level 2 conditions, relying exclusively on Ebola virus components.

Difficulty
advanced
Total time
~3–5 days (trVLP production ~2 days, infection and harvest ~1 day, assays ~1–2 days)
Model organism
Mammalian cell culture (species not specified)
Biosafety
BSL-2

Steps

1
Produce tetracistronic minigenome-containing virus-like particles

Generate replication and transcription-competent VLPs (trVLPs) using Ebola virus components in mammalian cell culture. This step establishes the safe BSL-2 compatible infectious particles for downstream lifecycle modeling.

▶ 03:18
2
Prepare target cells and infect with trVLPs

Culture and prepare mammalian target cells, then expose them to the previously produced trVLPs to initiate viral infection under controlled conditions.

▶ 05:41
3
Harvest infected cells and measure reporter activity

Collect infected cells at designated timepoints and quantify reporter activity to assess progression of the Ebola virus lifecycle across multiple replication cycles.

▶ 07:13
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