Quantitative PCR for counting residual white blood cells in blood products.
Leukocyte depleted blood components are frequently used to reduce alloimmunization and the risk of transfusion transmitted infection. Counting residual white blood cells in filtered blood products requires sensitive and reliable techniques. After separation of white blood cells from 500 microliters of 20 non-filtered and 54 filtered blood products we used polymerase chain reaction (PCR) and fluorimetric detection for the quantification of genomic DNA. The results were compared with results from Nageotte chamber counting. The accurate limit of detection of PCR was determined at 1 WBC/microliter (intra-assay coefficient of variation: 16.3%). PCR correlated well with Nageotte chamber counts (r = 0.77, p < 0.001, n = 74). Concordant results were obtained in 51 filtered and 20 non-filtered blood products. Discrepant results were obtained in 3 filtered whole blood units: In these blood products > 12 WBC/microliters were counted in Nageotte chamber and PCR gave a negative result. After component preparation fresh-frozen plasma and red cell concentrates of these units contained < 1 WBC/microliter using both methods. In conclusion we describe a quantitative PCR method which had about the same sensitivity and specificity as Nageotte chamber testing. However, PCR is more laborious than the standard method. As well, as reliable PCR testing requires expensive instruments and staff experienced in molecular biology, the standard method is more cost effective.
Legler
T J
TJ
Department of Transfusion Medicine, University of Göttingen, Germany. tleger@med.uni-goettingen.de
Riggert
J
J
Dove
G
G
Köhler
M
M
D005470:Fluorometry; D006801:Humans; D007958:Leukocyte Count / Q000379:methods*; D016133:Polymerase Chain Reaction / Q000379:methods*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity