HIV reservoir quantification by five-target multiplex droplet digital PCR

Area of Science:

• Virology
• Molecular Biology
• Immunology

Background:

• Most latent human immunodeficiency virus (HIV) proviruses are defective.
• Assessing the number of intact HIV proviruses is vital for evaluating HIV cure strategies.

Purpose of the Study:

• To develop and describe a method for quantifying potentially intact HIV proviruses.
• To establish a reliable clinical parameter for assessing HIV cure therapies.

Main Methods:

• High-molecular-weight DNA isolation followed by restriction enzyme fragmentation.
• Multiplexed droplet digital PCR to quantify five targets across the HIV genome.
• A reference assay to count T lymphocytes and assess DNA integrity.

Main Results:

• The described protocol allows for the estimation of potentially intact proviral copies.
• The method provides a quantitative measure relevant for clinical assessment of HIV cure.

Conclusions:

• This protocol offers a robust approach to measure intact HIV proviruses.
• Accurate quantification of intact HIV is a key step towards achieving an HIV cure.