Assessment of the effects of instrumentation, monoclonal antibody, and fluorochrome on flow cytometric immunophenotyping: a report based on 2 years of the NIAID DAIDS flow cytometry quality assessment program.
This study of the effect on CD4%, CD8%, CD3+8+%, and CD3% of flow cytometer, monoclonal antibody, and fluorochrome was based on 71 whole-blood samples, each evaluated by 42 to 59 laboratories during 2 years of a flow cytometry quality assessment program. For the 24 HIV-positive specimens, FACScans produced significantly lower CD4% values than EPICS-Cs or EPICS Profiles, and for the 47 HIV-negative specimens, FITC was associated with significantly lower CD4% values than PE or RD1, but differences were never larger than 2% and regressions accounted for only 3-12% of the variability. The labs using the most common CD4 technique had significantly higher between-laboratory variability than all other labs grouped together. For both CD8 and CD3+8+, measurements on FACScans were significantly higher than measurements on EPICS, and measurements using Leu2 were significantly higher than measurements using T8, with regressions accounting for 12-31% of the variability. The machine differences in medians were 3-7% for labs using Leu2-FITC. It might be worthwhile to discourage the use of Leu2-FITC for measuring CD8% but no change in instrument, monoclonal antibody, or fluorochrome would greatly improve interlaboratory agreement on CD4%.
Gelman
R
R
Harvard School of Public Health, Boston, Massachusetts 02115.
Cheng
S C
SC
Kidd
P
P
Waxdal
M
M
Kagan
J
J
D000911:Antibodies, Monoclonal / Q000276:immunology*; D015703:Antigens, CD / Q000032:analysis; D015214:Antigens, Differentiation, Myelomonocytic / Q000032:analysis; D017252:CD3 Complex / Q000032:analysis; D015704:CD4 Antigens / Q000032:analysis; D016827:CD8 Antigens / Q000032:analysis; D002469:Cell Separation; D005434:Flow Cytometry / Q000295:instrumentation; D005456:Fluorescent Dyes; D006679:HIV Seropositivity / Q000276:immunology*; D006801:Humans; D016130:Immunophenotyping / Q000592:standards*; D017493:Leukocyte Common Antigens / Q000032:analysis; D018950:Lipopolysaccharide Receptors
D000911:Antibodies, Monoclonal; D015703:Antigens, CD; D015214:Antigens, Differentiation, Myelomonocytic; D017252:CD3 Complex; D015704:CD4 Antigens; D016827:CD8 Antigens; D005456:Fluorescent Dyes; D018950:Lipopolysaccharide Receptors; D017493:Leukocyte Common Antigens