QUANTITATION OF CD38 EXPRESSION USING QUANTIBRITE(TM) BEADS

The QuantiBRITE(TM) bead method was compared with the CD4 biological c alibration method for quantitation of CD38 expression on CD8(+) T-lymp hocytes of Multicenter AIDS Cohort Study participants. Results were ex pressed as CD38 antibodies bound per cell (ABCs) and were the same wit h the two methods provided two conditions were met. These were the use of repurified (> 95% of the monoclonal antibodies [mAbs] hare 1 phyco erythrin [PE] molecule per mAb) CD38-PE for both methods and use of re purified CD4-PE to calculate the relative fluorescence intensity multi plier for the CD4 biological calibration method. Our results indicate that the prognostic significance of CD38 values obtained using the Qua ntiBRITE method can be interpreted using previously published reports (Liu et al.: J Acquir Immune Defic Syndr Hum Retrovirol 16:83-92, 1997 and 18:332-340, 1998). Sample preparation using NH4Cl and FAGS lysing solution gave similar results for CD38 relative fluorescence intensit y. Dilution into either phosphate-buffered saline with 2% fetal calf s erum and 0.1% sodium azide or fixation in 1% paraformaldehyde for 1 or 24 h also gave similar results. In experiments using Raji cells, whic h express high levels of CD38, the valence of binding of the intact Le u 17 antibody was similar to 68% bivalent and similar to 32% monovalen t. This emphasizes the complexity of determining antigen density from ABCs. We conclude that repurified PE conjugates of CD38, which can be consistently made, together with QuantiBRITE PE beads, provide a conve nient and reliable method for quantitation of CD38 expression as ABCs. (C) 1998 Wiley-Liss, Inc.