Collection and Isolation of CD14+ Primary Human Monocytes Via Dual Density Gradient Centrifugation as a Model System to Study Human Cytomegalovirus Infection and Pathogenesis.

Human cytomegalovirus (HCMV) can cause severe disease in the immunocompromised. One of the hallmarks of HCMV infection of a human host is the targeted infection of peripheral blood monocytes (but not other leukocyte populations) that in turn serve as the key cell type for hematogenous dissemination and the establishment of persistence following primary infection. Monocytes are also a key cell type associated with viral reactivation and spread following viral reactivation. Because of their importance in the HCMV-host infection cycle and lifelong infection, it is critical to be able to study their infection in controlled in vitro systems in the laboratory. In this chapter, we discuss a viable protocol for harvesting fresh ex vivo blood monocytes from human donors that are pure and unactivated cells and that can be used in a research setting.