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Comparison of five platforms for enumeration of residual leucocytes in leucoreduced blood components
P F van der Meer1, J W Gratama, C J van Delden
1Blood Bank North Holland, Pkesmanlaan 125, 1066 CX Amsterdam, The Netherlands. bnhams@hetnet.nl
The need for quality control of leucoreduction of blood products has led to the development of various methods to count low levels of residual leucocytes. We compared five platforms side-by-side: the Nageotte haemocytometer and four based on fluorescent staining of nuclei: two flowcytometers (Beckman Coulter, BD Biosciences) with methods based on counting beads, a volumetric flow cytometer (Partec) and the microvolumic fluorimeter ImagN2000 (BD Biosciences), all according to their manufacturers' recommended methods. Analysis of double-filtered red cell concentrates (RCCs) and platelet concentrates (PCs), spiked with various numbers of leucocytes, revealed good linearity for all methods over the range of 1.6-32.7 leucocytes/microl, all with r(2) > 0.99. At the rejection level of leucocyte-reduced blood components, i.e. 1 x 10(6) per unit corresponding with approximately 3.3 leucocytes/microl, the Nageotte haemocytometer had low accuracy (0% for RCCs, 56% for PCs), and was relatively imprecise [coefficient of variance (CV) of 34% and 30% respectively]. The Partec flow cytometer gave good results for RCCs (accuracy 67%, CV 22%), but not for PCs (accuracy 0%, CV 25%). The ImagN2000 had an accuracy of 44% for RCCs and 89% for PCs, but the precision was variable (CV 32% for RCCs, 15% for PCs). The best results were obtained with the Beckman Coulter (RCCs: accuracy 86%, CV 13%, PCs: accuracy 67%, CV 16%), and BD Biosciences platforms (RCCs: accuracy 100%, CV 10%; PCs: accuracy 89%, CV 11%). We conclude that, at the rejection level of 1 x 10(6) leucocytes per unit, the widely used Nageotte haemocytometer performs poorly in terms of inaccuracy and imprecision, and that both counting-bead-based, flow cytometric methods performed best.
The need for quality control of leucoreduction of blood products has led to the development of various methods to count low levels of residual leucocytes. We compared five platforms side-by-side: the Nageotte haemocytometer and four based on fluorescent staining of nuclei: two flowcytometers (Beckman Coulter, BD Biosciences) with methods based on counting beads, a volumetric flow cytometer (Partec) and the microvolumic fluorimeter ImagN2000 (BD Biosciences), all according to their manufacturers' recommended methods. Analysis of double-filtered red cell concentrates (RCCs) and platelet concentrates (PCs), spiked with various numbers of leucocytes, revealed good linearity for all methods over the range of 1.6-32.7 leucocytes/microl, all with r(2) > 0.99. At the rejection level of leucocyte-reduced blood components, i.e. 1 x 10(6) per unit corresponding with approximately 3.3 leucocytes/microl, the Nageotte haemocytometer had low accuracy (0% for RCCs, 56% for PCs), and was relatively imprecise [coefficient of variance (CV) of 34% and 30% respectively]. The Partec flow cytometer gave good results for RCCs (accuracy 67%, CV 22%), but not for PCs (accuracy 0%, CV 25%). The ImagN2000 had an accuracy of 44% for RCCs and 89% for PCs, but the precision was variable (CV 32% for RCCs, 15% for PCs). The best results were obtained with the Beckman Coulter (RCCs: accuracy 86%, CV 13%, PCs: accuracy 67%, CV 16%), and BD Biosciences platforms (RCCs: accuracy 100%, CV 10%; PCs: accuracy 89%, CV 11%). We conclude that, at the rejection level of 1 x 10(6) leucocytes per unit, the widely used Nageotte haemocytometer performs poorly in terms of inaccuracy and imprecision, and that both counting-bead-based, flow cytometric methods performed best.