The clinical usefulness of monitoring CD4+ T-lymphocyte counts in patients infected with HIV is now well established. The need for accurate, rapid, and cost-effective methods for making these determinations is evident. Recent technologic advances have allowed for the development of a system for the determination of CD4+ T-lymphocyte counts by simultaneous 4-color flow cytometry. A new 4-color 2-tube flow cytometric method for analyzing CD4+ T-lymphocyte subsets in whole blood was compared with a standard 2-color 5-tube method. The new method provides results almost identical to those of the well-established 2-color method used in our clinical laboratory. Statistical analyses indicate very low variability in CD4+ counts between the 2 methods, strongly supporting the usefulness of this new procedure. In addition, the 4-color procedure provides a 15% reduction in the materials cost per test compared with the 2-color method, as well as a marked reduction in the time expenditure of flow cytometry technologists.