Intra- and interlaboratory variability of paroxysmal nocturnal hemoglobinuria testing by flow cytometry following the 2012 Practical Guidelines for high-sensitivity paroxysmal nocturnal hemoglobinuria testing
Article first published online: 2 AUG 2013
Copyright © 2013 International Clinical Cytometry Society
Cytometry Part B: Clinical Cytometry
Volume 84B, Issue 4, pages 229–236, July/August 2013
How to Cite
How to cite this article: Intra- and interlaboratory variability of paroxysmal nocturnal hemoglobinuria testing by flow cytometry following the 2012 practical guidelines for high-sensitivity paroxysmal nocturnal hemoglobinuria testing. Cytometry Part B 2013; 84B: 229–236., , , , , , , , .
- Issue published online: 2 AUG 2013
- Article first published online: 2 AUG 2013
- Manuscript Accepted: 19 DEC 2012
- Manuscript Revised: 6 DEC 2012
- Manuscript Received: 22 OCT 2012
- Alexion Pharma International
- flow cytometry;
- ICCS Guidelines;
- intra and interlaboratory variability
Sutherland et al. recently published the Practical Guidelines for high-sensitivity detection of paroxysmal nocturnal hemoglobinuria (PNH) clones by flow cytometry (FCM), containing concise protocols for PNH testing.
Using this approach, we studied the intra- and interlaboratory variability observed in a multicenter study in which fresh blood samples containing three clinically relevant PNH clone sizes within the granulocytic, monocytic, and red blood cell (RBC) populations were shipped to each participating center.
Coefficients of variation (CVs) for precision/reproducibility analysis ranged from 0.01%/0.02% to 0.48%/0.45% (big clone), from 0.69%/1.52% to 4.24%/5.80% (small-intermediate clone), from 1.47%/3.91% to 15.01% /17.83% (minor clone) for PNH white blood cells (WBCs) and from 0.24%/0.48% to 1.76%/1.83% (big clone), from 0.80%/1.14% to 2.39%/4.45% (small-intermediate clone), from 1.09%/3.36% to 10.54%/10.23% (minor clone) for PNH RBCs, respectively. Linear regression analysis showed excellent performance correlation between centers (r > 0.99), Wilcoxon rank test revealed no statistically significant differences for PNH granulocytes, monocytes, and RBCs (P > 0.05%), Bland–Altman analysis demonstrated good performance agreement for all target PNH clones (mean bias ranging from −1.47 to 0.71).
Our results demonstrate very good intra- and interlaboratory performance characteristics for both precision and reproducibility analyses and excellent correlation and agreement between centers for all target PNH clone sizes. Our data confirm the reliability and robustness of the recently published Practical Guidelines approach for high sensitivity PNH testing by flow cytometry and suggest that such an approach represents an excellent basis for standardization of PNH testing by flow cytometry. © 2013 International Clinical Cytometry Society