Use of formalin-fixed, propidium iodide-stained human leukocytes as a standard for enumerating CD4(+) T lymphocytes in a single-platform assay

A new, inexpensive method is described that enables lymphocytes to be enume rated very precisely. Normal leukocytes were simultaneously stained and fix ed with a propidium iodide-paraformaldehyde solution. The preparation obtai ned (CellBeads) was used as an internal standard for cell enumeration by fl ow cytometry and was stable at 4 degreesC for at least 60 days. Unlike synt hetic beads, the CellBeads behaved similarly to normal cells during red blo od cell lysis and cell mashing procedures, When known numbers of CellBeads were added to whole blood samples and the numbers of CellBeads and lymphocy tes were determined, highly reproducible and accurate enumerations were obt ained-far more so than when synthetic beads were used. This inexpensive met hod is suitable for routine use.