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A flow cytometric method for measuring neutralization of HIV-1 subtype B and E primary isolates
J M Darden1, V R Polonis, M S deSouza
1Office for Research and Development, Center of Excellence for Flow Cytometry, Siriraj Hospital, Mahidol University, Bangkok, Thailand.
Cytometry
|May 11, 2000
Summary
This study introduces flow cytometry as a reliable method for HIV-1 neutralization assays. It accurately measures responses to diverse HIV-1 subtypes, improving vaccine development.
Area of Science:
- Virology
- Immunology
- Vaccine Development
Background:
- Clinical trials for human immunodeficiency virus type 1 (HIV-1) vaccines rely on neutralization assays.
- Current p24 assays show variable results for different HIV-1 subtypes.
- Flow cytometry offers a potential alternative for monitoring HIV-1 neutralization.
Purpose of the Study:
- To evaluate flow cytometry's utility in assessing HIV-1 primary isolate neutralization.
- To establish a more reliable endpoint for HIV neutralization assays.
Main Methods:
- A modified neutralization assay using CD8-depleted peripheral blood mononuclear cells (PBMC).
- Cells were stained for intracellular HIV-1 p24 and analyzed via flow cytometry.
- HIV-1 subtypes B' and E primary isolates were tested with pooled patient sera/plasma.
Main Results:
- Uninfected cells showed <0.2% p24(+) cells.
- Infected cultures without antibody had 18-42% p24(+) cells.
- Subtype-specific neutralization ranged from 0% to 99% reduction in infected cells.
Conclusions:
- Flow cytometry for intracellular HIV-1 p24 detection is a viable endpoint assay.
- This method allows for early identification of p24 in specific cell subsets.
- It provides a quantitative and alternative endpoint for HIV neutralization assays.