High-sensitivity 5-, 6-, and 7-color PNH WBC assays for both Canto II and Navios platforms
Language English Country United States Media print-electronic
Document type Journal Article
PubMed
29381839
DOI
10.1002/cyto.b.21626
Knihovny.cz E-resources
- Keywords
- cross-platform, high sensitivity, paroxysmal nocturnal hemoglobinuria, white blood cells,
- MeSH
- Antigens, CD analysis MeSH
- Immunophenotyping instrumentation methods MeSH
- Humans MeSH
- Hemoglobinuria, Paroxysmal diagnosis MeSH
- Flow Cytometry instrumentation methods MeSH
- Sensitivity and Specificity MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
- Names of Substances
- Antigens, CD MeSH
BACKGROUND: Paroxysmal Nocturnal Hemoglobinuria (PNH) is a rare acquired hematopoietic stem cell disorder characterized by an inability to make Glyco-Phosphatidyl-Inositol (GPI)-linked cell surface structures. Fluorescent proaerolysin (FLAER-Alexa488) is increasingly used to detect GPI-deficient WBCs by flow cytometry. However, FLAER is not available in all countries and is expensive to obtain in others. An earlier study to compare FLAER-based and non-FLAER assays confirmed very good agreement between the two tubes suggesting a cost effective simultaneous evaluation of PNH neutrophils and monocytes is possible without FLAER. METHODS: We have used a single tube approach with a 7-color assay comprising FLAER-CD157-CD15-CD64-CD24-CD14-CD45. Conjugates were carefully selected and validated so that stained samples could be analyzed on either 10-color Navios or 8-color FACSCanto II platforms. The 6-color (minus CD14) and 5-color (minus CD24 and CD14) versions were also developed and compared with our predicate clinical lab 5-color assay comprising FLAER-CD157PE-CD64ECD-CD15PC5-CD45PC7. RESULTS/CONCLUSIONS: CD15-gated PNH neutrophil clone size was quantified using either FLAER and CD157, FLAER and CD24, or CD157 and CD24. CD64-gated PNH monocyte clone size was quantified using either FLAER and CD157, FLAER and CD14, or CD157 and CD14. Analysis of >40 PNH samples showed that the FLAER-based plots derive virtually identical data to the non-FLAER plot for neutrophils (R2 = 1) and monocytes (R2 = 0.9999) and that closely similar data can be acquired using both Canto II and Navios platforms with 7-, 6-, and 5-color versions of the assay. Assessment of non-PNH samples confirmed extremely low background rate of PNH phenotypes (neutrophils and monocytes) with all three approaches. © 2018 International Clinical Cytometry Society.
Cytoquest Corporation Toronto Ontario Canada
Dahl Chase Diagnostic Services Bangor Maine
EXBIO Praha a s Vestec Czech Republic
Institute of Hematology and Blood Transfusion Prague Czech Republic
Laboratory Medicine Program Toronto General Hospital Ontario Canada
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