Home › Cell Biology › Visualizing Cell-to-cell Transfer of HIV using Fluorescent Clones of HIV and Live Confocal Microscopy
Cell Biology JoVE (Open Access) Citable · DOI

Visualizing Cell-to-cell Transfer of HIV using Fluorescent Clones of HIV and Live Confocal Microscopy

DOI: 10.3791/2061-v
What you'll learn
  • ✓ Set up and prepare donor and target CD4+ T cells for HIV transfer visualization
  • ✓ Label and monitor live cell-to-cell HIV transfer using fluorescent molecular clones
  • ✓ Acquire and interpret confocal microscopy images of HIV Gag-iGFP dynamics
Protocol

This visualized experiment is a guide for utilizing a fluorescent molecular clone of HIV for live confocal imaging experiments.

Difficulty
advanced
Total time
~4–6 hours (cell preparation, labeling, and live imaging)
Model organism
Jurkat T cells (human T-cell line), Primary human CD4+ T cells
Biosafety
BSL-2

Steps

1
Prepare donor and target CD4+ T cells

Culture and harvest Jurkat donor cells and primary human CD4+ T cells. Prepare cells at appropriate densities for co-culture and imaging.

▶ 01:26
2
Clean donor cells and label target cells

Remove media and debris from donor cells; apply fluorescent labeling to target CD4+ T cells to enable discrimination during live-cell imaging.

▶ 03:47
3
Monitor HIV cell-to-cell transfer via confocal microscopy

Set up live confocal imaging of co-cultured cells and acquire time-lapse sequences to visualize HIV Gag-iGFP transfer from donor to target cells.

▶ 06:36
4
Analyze representative images of HIV transfer

Review and interpret confocal images showing HIV Gag-iGFP accumulation and cell-to-cell transfer events.

▶ 12:02
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