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Related Experiment Video

Updated: Jun 22, 2025

Flow Virometry to Analyze Antigenic Spectra of Virions and Extracellular Vesicles
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Applying Flow Virometry to Study the HIV Envelope Glycoprotein and Differences Across HIV Model Systems.

Jonathan Burnie1,2,3, Claire Fernandes1,2, Ayushi Patel1,2

  • 1Department of Biological Sciences, University of Toronto Scarborough, Toronto, ON M1C 1A4, Canada.

Viruses
|June 27, 2024
PubMed
Summary

Flow virometry effectively characterizes the HIV envelope glycoprotein (Env) on single virions. This method reveals Env heterogeneity and conformational changes, aiding HIV vaccine development.

Keywords:
Env conformationHIV EnvHIV trimercalibrated flow virometrygp120/gp41human immunodeficiency virus (HIV)molecules of equivalent soluble fluorophore (MESF)nanoscale flow cytometryneutralizationvirion capture

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Area of Science:

  • Virology
  • Immunology
  • Biochemistry

Background:

  • The HIV envelope glycoprotein (Env) is crucial for viral entry and a key target for immune responses and vaccine design.
  • Challenges in targeting Env include its glycosylation, variability, conformational masking, and low abundance on virions.
  • Understanding Env's structure and presentation is vital for effective therapeutic and preventative strategies.

Purpose of the Study:

  • To demonstrate the utility of flow virometry (FV) for high-throughput, single-virion phenotyping of the HIV Env.
  • To characterize Env heterogeneity and conformational states using a comprehensive antibody panel.
  • To assess the impact of virus-producing cell models on Env detection.

Main Methods:

  • Staining of HIV virions with 85 monoclonal antibodies targeting various Env regions.
  • Application of flow virometry for quantitative analysis of single virion Env expression.
  • Comparison of Env detection on viruses produced from different cell lines (CD4+ T cells, HEK293T, primary cells).
  • Assessment of Env conformational changes upon addition of soluble CD4.

Main Results:

  • A wide range of antibodies successfully stained HIV Env, with V3 antibodies showing the highest signal.
  • The cellular model used for virus production significantly influenced Env detection.
  • Flow virometry detected heterogeneity in Env presentation on individual virions.
  • FV sensitively identified conformational differences in Env upon soluble CD4 binding.

Conclusions:

  • Flow virometry is a powerful tool for detailed characterization of HIV Env at the single-virion level.
  • FV can reveal critical insights into Env heterogeneity and conformational dynamics, relevant for vaccine design.
  • The choice of cell model impacts Env detection, highlighting the need for relevant systems in research.