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- Title
ORIGINAL PAPER Influence of cell-free DNA in plasma on real-time polymerase chain reaction for determination of residual leucocytes in platelet concentrates.
- Authors
Dijkstra-Tiekstra, M. J.; Pietersz, R. N. I.; Reesink, H. W.; van der Schoot, C. E.
- Abstract
Real-time quantitative (RQ) polymerase chain reaction (PCR) can be used to determine the number of residual leucocytes in leucocyte-reduced platelet concentrates (LR-PCs), which should contain < 3·3 leucocytes/µl. In this study we investigated the extent to which cell-free DNA, known to be present in plasma, might interfere with this determination. In this study, RQ-PCR was employed to determine the following: the influence of filtration of platelet concentrates (PCs) on the amount of cell-free DNA; the variation in concentration of cell-free DNA between the buffy coats (BCs) of different donors; and the amount of cell-free DNA during storage and processing of whole blood. PCs were sampled before and after filtration ( n = 5), BCs were sampled ( n = 100) and whole blood units were sampled < 2 h and 16–20 h after collection, and the BCs were also sampled after processing the whole blood ( n = 10). Samples were centrifuged to obtain cell-free plasma in which the amount of cell-free DNA was determined using an RQ-PCR for the albumin gene. The amount of cell-free DNA was not influenced by filtration of the PCs [1·7 ± 0·8 vs. 1·5 ± 0·8 leucocyte-equivalents (eq)/µl]. However, the amount of cell-free DNA in plasma of the BCs varied considerably, from 0·1 to 18·2 leucocyte-eq/µl (median = 1·5 leucocyte-eq/µl; mean ± SD: 2·2 ± 2·4 leucocyte-eq/µl). In 18% of the BCs the amount cell-free DNA was > 3·3 leucocyte-eq/µl. The amount of cell-free DNA increased during storage, from 0·3 ± 0·3 leucocyte-eq/µl (< 2 h after collection) to 0·9 ± 0·6 leucocyte-eq/µl (16–20 h after collection) and, after processing the whole blood, to 2·0 ± 2·0 leucocyte-eq/µl. Variable amounts of cell-free DNA in plasma will interfere if RQ-PCR is applied to estimate leucocyte numbers in leucocyte-reduced PCs.
- Subjects
DNA; BLOOD plasma; LEUCOCYTES; POLYMERASE chain reaction; BLOOD platelets; NUCLEIC acids
- Publication
Vox Sanguinis, 2004, Vol 86, Issue 2, p130
- ISSN
0042-9007
- Publication type
Article
- DOI
10.1111/j.0042-9007.2004.00402.x