Isolation of human immunodeficiency virus type 1 from peripheral blood monocytes

Human immunodeficiency virus type 1 (HIV-1) can infect circulating peripheral blood monocytes and resting CD4+ T lymphocytes despite sustained suppression of plasma viremia to undetectable levels. These persistently infected cell populations pose a barrier for virus eradication by highly active antiretroviral therapy (HAART), and are a significant reservoir of HIV-1 contributing to viral rebound following cessation or failure of HAART. This chapter provides a protocol for isolating replication-competent HIV-1 from peripheral blood monocytes of HIV-1-infected individuals, including those with sustained plasma HIV-1 RNA levels below 50 copies/mL, by co-culture with CD8-depleted, phytohemagglutinin-activated donor peripheral blood mononuclear cells. In our laboratory, this protocol has the sensitivity to achieve a success rate of positive HIV-1 isolation in approx 70% of cases. The study of HIV-1 strains harbored by peripheral blood monocytes of patients undergoing HAART will contribute to the understanding of viral persistence in cellular reservoirs that impede effective HAART.