Abstract
Background: In the past decade, human immunodeficiency virus (HIV) lymphocyte immunophenotyping has evolved significantly. New fluorochromes, new multicolor reagents, enhanced instruments, and the capacity to provide absolute cell counts using the single-platform technique have all contributed to the reliability of T-cell subset measurements. In this study, four gating protocols were evaluated to select the most robust method for T-cell subset enumeration. Methods: Peripheral blood specimens from 21 HIV+ and 20 HIV - individuals were monitored up to 96 h. Aliquots of specimens were stored at room temperature and analyzed at 6 (baseline), 48, 72, and 96 h. Aliquots were stained with CD45-fluorescein isothtocyanate (FITC)/CD3PC5/CD4RD1/ CD8ECD. Data analysis was performed with all four gating protocols. Results: Only with fresh blood did all protocols provide similar results. From samples that were 48 h old, the choice of gating strategy had a dramatic impact on immunophenotyping results. The largest deviations from baseline values occurred at 96 h and gating protocols that included dual light scatter gates provided the greatest shift of T-cell subset values over time. The gating protocols that were based exclusively on cell lineage-specific gates gave the most robust T-cell values up to 96 h. Conclusion: By selecting the appropriate gating protocol, the temporal integrity of specimens can be extended up to 4 days.
Original language | English (US) |
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Pages (from-to) | 53-61 |
Number of pages | 9 |
Journal | Clinical Cytometry |
Volume | 50 |
Issue number | 2 |
DOIs | |
State | Published - 2002 |
Keywords
- CD45 gating
- Double anchor gate
- Four-color immunophenotyping
- Homogeneous and heterogeneous gating parameters
- Integrated primary gate
- Intrinsic and extrinsic parameters
- Light scatter gate
- Sequential mixed gate
- Single-platform absolute counting
- T gating
ASJC Scopus subject areas
- Pathology and Forensic Medicine
- Histology
- Cell Biology