First published online July 26, 2007
Stem Cells
Vol. 25 No.
11
November 2007, pp.
2972
doi:10.1634/stemcells.2007-0519; www.StemCells.com
© 2007 AlphaMed Press
Beyond the FACS
Raymond Barfielda,
Xiaohua Chena,
Mario Ottoa,
Rupert Handgretingerb
aSt. Jude Children's Research Hospital, Hematology/Oncology, Memphis, Tennessee, USA;
bUniversity of Tubingen, Pediatrics, Tubingen, Germany
Key Words. CD34+ stem cells • CD34– hematopoietic stem cells • Hematopoietic stem cell transplantation
Correspondence: Raymond Barfield, M.D., Ph.D., St. Jude Children's Research Hospital, Hematology/Oncology, Memphis, Tennessee 38105, USA. Telephone: 901-495-4720; Fax: 901-521-9005; e-mail: raymond.barfield{at}stjude.org
Received July 2, 2007;
accepted for publication July 20, 2007.
First published online in STEM CELLS EXPRESS July 26, 2007.
We appreciate the thoughtful response from Robert Sutherland and Michael Keeney [1]. We found their review of the complexities of flow cytometry involving various anti-CD34 antibodies interesting and succinct. However, we are not at all sure that their comments are at odds with our findings. Our primary point was that by selecting two single CD34+ populations with different approaches, significant differences in signal-joint T-cell receptor excision circle (sjTREC) production were observed. If this is the case (and we state explicitly that further experiments are warranted to confirm or disconfirm these findings), there may well be useful stem cells lost through approaches to graft engineering that make use of a single antibody targeting a single stem cell-associated antigen. We make no claim whatsoever that the use of the QBEnd has not been successful in the clinical setting. That is to say, our aim is not to "fly in the face" of past success but rather to offer preliminary data suggesting that alternative approaches might improve this successful track record. For example, negative selection in which CD3+ cells are removed from the graft (as opposed to positive selection in which CD34+ cells are selected) allows retention in the graft not only of CD34+ stem cells that might be lost using a single anti-CD34 antibody for selection but also CD34– stem cells. Anyone who has been involved in CD34+ stem cell transplantation knows that there is room for improvement, especially perhaps in T-cell reconstitution.
It may be that the emphasis placed by Sutherland and Keeney on the details of cytometry rather than on the data we present regarding differences in sjTREC production between two populations of stem cells follows from the fact that we included data on two other antibodies, 12.8 and 581. In retrospect, we agree that these flow data do not add to the central point of the paper and indeed may be more of a distraction. However, this does not bear on the fundamental points we make. To emphasize again, we take our data as preliminary but suggestive, and indeed we have already begun using CD3 and CD3/CD19 depletion strategies in our institutions precisely to avoid the potential loss of CD34+ stem cells that might be missed by a single antibody as well as CD34– stem cells that may also contribute to faster immune recovery—our ultimate goal.
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DISCLOSURE OF POTENTIAL CONFLICTS OF INTEREST
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The authors indicate no potential conflicts of interest.
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REFERENCES
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- Sutherland DR, Keeney M. Re: Selection of stem cells by using antibodies that target different CD34 epitopes yields different patterns of T-cell differentiation. STEM CELLS 2007;25:2385–2386.[Free Full Text]
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