Recent interest focused on the dynamics of HIV-1 replication in primary monocytes/macrophages and T-lymphocytes of the immune system, as well as the standardization of virological and immunologicalin vitroassays with primary isolates, provided the impetus for these studies. These types of studies have never been performed as they would occurin vivo,i.e., where the envelope of the virus and cell membranes of the two cell types of the same host origin. Therefore, the biological and physicochemical properties of an uncloned, primary dual-tropic isolate HIV-1_ADAduring the initial lag, log, and stationary phases of viral replication were studied in an autologous donor cell assay in peripheral blood mononuclear cells (PBMC) and blood monocyte-derived macrophages (MDM). Similar total numbers (10⁹virus particles/ml) were produced by both cell types during the stationary period. On a per cell per day basis, during peak stationary periods, 0.92 × 10³virions/day for MDMs and 5.31 × 10³virions/day for PBMCs were produced. Interestingly, virus replicating from MDMs during the log-growth phase demonstrated the greatest infectious fraction which was 3 logs greater than virus replicating in PBMCs. Despite constant virus particle production in MDMs, the infectious fraction was found to fall 3 to 4 logs over a 10-day period. Due to an infectious fraction less than 1 (0.053 infectious unit/cell/24 hr), virus spread in PBMCs during the rapid log phase could only have occurred by cell-to-cell contact, whereas in MDMs with an infectious fraction of about one infectious particle (∼1/cell/24 hr), cell-free transmission could account for the observed results. Most of the MDMs (>90%) became productively infected, whereas only 5–10% of the total PBMCs were found replicating virus. The period of peak stationary virus production (i.e., stationary phase) was at minimum 4 to 5 times longer in MDMs than PBMCs. Whereas the majority of p24, RT, and gp120 found to be associated with MDM-derived virions, no increased dissociation of these components was observed in PBMC-derived virions. The virion-associated gp120 was 3 to 4 times more stable on both PBMC- and MDM-derived virus (>96 hr) and present at 10–25 times the concentration per virion than that observed for a T-cell-line-adapted laboratory strain of HIV-1 replicating in T-cell lines. Thesein vitroresults suggest that important differences exist between MDMs and PBMCs with regard to the viral dynamics of infection and replication which should provide for a qualitative and quantitative basis to estimate virus replication on a per-cell basis for other known cellular targets of HIV-1. Studying the multiple biophysicochemical characteristics and viral replication dynamics as described herein provides an autologousin vitromodel of additional quantifiable parameters for analysis and understanding of virus/host factor(s) and/or antivirals which influence them.