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Discrimination of Seven Immune Cell Subsets by Two-fluorochrome Flow Cytometry
Published on: March 5, 2019
Monitoring temporary immunodepression by flow cytometric measurement of monocytic HLA-DR expression: a multicenter
Wolf-Dietrich Döcke1, Conny Höflich, Kenneth A Davis
1Institute of Medical Immunology, Charité-Campus Mitte, Humboldt University of Berlin, and University Hospital, Charité Berlin, Germany.
Background:
Single-center trials have shown that monocytic HLA-DR is a good marker for monitoring the severity of temporary immunodepression after trauma, major surgery, or sepsis. A new test for measuring monocytic HLA-DR is now available.
Methods:
We evaluated a new test reagent set for monocytic HLA-DR expression (BD Quantibritetrade mark HLA-DR/Monocyte reagent; Becton Dickinson) in single-laboratory and interlaboratory experiments, assessing preanalytical handling, lyse-no-wash (LNW) vs lyse-wash (LW) values, reference values, and the effect of use of different flow cytometers and different instrument settings on test variance.
Results:
For preanalytical handling, EDTA anticoagulation, storage on ice as soon as possible, and staining within 4 h after blood collection gave results comparable to values obtained for samples analyzed immediately after collection (mean increase of approximately 4% in monocytic HLA-DR). Comparison of LNW and LW revealed slightly higher results for LNW ( approximately 18% higher for LNW compared with LW; r = 0.982). Comparison of different flow cytometers and instrument settings gave CVs <4%, demonstrating the independence of the test from these variables and suggesting that this method qualifies as a standardized test. CV values from the interlaboratory comparison ranged from 15% (blood sample unprocessed before transport) to 25% (stained and fixed before transport).
Conclusions:
For the BD Quantibrite HLA-DR/Monocyte test, preanalytical handling is standardized. Single-laboratory results demonstrated the independence of this test from flow cytometer and instrument settings. Interlaboratory results showed greater variance than single-laboratory values. This interlaboratory variance was partly attributable to the influence of transport and can be reduced by optimization of transport conditions.
