Normal human blood density gradient lymphocyte subset analysis: I. An interlaboratory flow cytometric comparison of 85 normal adults

Abstract

An interlaboratory flow cytometric comparison of several commercially available human lymphocyte subset reagents was undertaken in three different laboratories. Fresh Hypaque‐Ficoll purified blood mononuclear cells were stained at 4°C or 22°C. Direct or indirect surface immunofluorescence was carried out at all sites using an EPICS V flow cytometer. Fullbright, 10‐μm fluorescent polystyrene microspheres were used for optical alignment and standardization. A log integral fluorescent histogram gated on forward and right angle scatter was collected on 1–2 × 10⁴ cells for each reagent and the proportion, of positive cell determined for each reagent. With the exception of one reagent, anti‐B1, which showed an approximately twofold variation, all three laboratories showed remarkable agreement. Thus there was no significant difference noted for the following reagents: OKT4, CCT4, Leu 3a, Leu 2a, OKT8, or CCT8. We attribute these findings to the availability of quality reagents, precision instrumentation, and a standard lymphocyte preparation.