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Stem Cells International
Volume 2013 (2013), Article ID 823912, 10 pages
http://dx.doi.org/10.1155/2013/823912
Research Article

The Assessment of Parameters Affecting the Quality of Cord Blood by the Appliance of the Annexin V Staining Method and Correlation with CFU Assays

1Institute of Transplantation Diagnostics and Cell Therapeutics, Heinrich Heine University Medical Center, Moorenstraße 5, 40225 Düsseldorf, Germany
2Anthony Nolan Research Institute, Royal Free Hospital, Fleet Road, London NW3 2QG, UK
3Department of Haematology, UCL Cancer Institute, Royal Free Campus, Pond Street, London NW3 2QG, UK
4Cord Blood Bank, Haematology Department, Careggi University Hospital, Via delle Oblate 1, 50141 Firenze, Italy
5Eurocord ARTM, Carré Historique Porte 05, Hôpital Saint-Louis, 1 Avenue Claude Vellefaux, 75010 Paris, France
6Barcelona CBB, Banc de Sang i Teixits, Passeig Taulat 116, 08005 Barcelona, Spain

Received 8 October 2012; Accepted 28 December 2012

Academic Editor: David Allan

Copyright © 2013 Teja Falk Radke et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

The assessment of nonviable haematopoietic cells by Annexin V staining method in flow cytometry has recently been published by Duggleby et al. Resulting in a better correlation with the observed colony formation in methylcellulose assays than the standard ISHAGE protocol, it presents a promising method to predict cord blood potency. Herein, we applied this method for examining the parameters during processing which potentially could affect cord blood viability. We could verify that the current standards regarding time and temperature are sufficient, since no significant difference was observed within 48 hours or in storage at up to . However, the addition of DMSO for cryopreservation alone leads to an inevitable increase in nonviable haematopoietic stem cells from initially 14.8% ± 4.3% to at least 30.6% ± 5.5%. Furthermore, CFU-assays with varied seeding density were performed in order to evaluate the applicability as a quantitative method. The results revealed that only in a narrow range reproducible clonogenic efficiency (ClonE) could be assessed, giving at least a semiquantitative estimation. We conclude that both Annexin V staining method and CFU-assays with defined seeding density are reliable means leading to a better prediction of the final potency. Especially Annexin V, due to its fast readout, is a practical tool for examining and optimising specific steps in processing, while CFU-assays add a functional confirmation.