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Development of a simplified counterflow centrifugation elutriation procedure for depletion of lymphocytes from human bone marrow. Transplantation 1986 Feb;41(2):220-9

Date

02/01/1986

Pubmed ID

2935979

DOI

10.1097/00007890-198602000-00017

Scopus ID

2-s2.0-0022650754   35 Citations

Abstract

Graft-versus-host disease (GVHD) remains a major complication of allogeneic bone marrow (BM) transplantation. Techniques that effectively purge BM of mature T lymphocytes should reduce the incidence of GVHD and improve survival. We have developed a simplified, two-flow rate, fixed rotor speed counterflow centrifugation-elutriation (CCE) procedure that reproducibly depletes 99% of lymphocytes from Ficoll-Hypaque(F/H)-separated BM or BM buffy-coat. Two predetermined flow rates (24 and 28 ml/min) were used to purge small and intermediate-to-large lymphocytes, respectively, whereas faster sedimenting cells were recovered at the termination of the run. Lymphocyte depletion was substantiated by pan-T monoclonal antibody analysis as well as by complete loss of responsiveness to alloantigens and mitogens. Despite the lack of mature T cells, the depleted marrow fraction retained lymphoid colony-forming ability. Lymphocyte-purged marrow was obtained in high yield (72%), and retained high viability (greater than 97%) and hematopoietic colony-forming ability (greater than 99%). The ratio of total myeloid/erythroid colony-forming cells to T lymphocytes was 73-fold higher in the lymphocyte-depleted fraction than in unseparated BM. We concluded that a two-step CCE procedure can be used to rapidly deplete lymphocytes from both F/H-separated and buffy-coat BM inocula without altering hematopoietic capacity as measured by the in vitro clonogenic assays. It may be possible to adapt this procedure to the separation of the large number of marrow cells required for human BM transplantation.

Author List

Noga SJ, Donnenberg AD, Schwartz CL, Strauss LC, Civin CI, Santos GW

Author

Cindy L. Schwartz MD, MPH Chief, Professor in the Pediatrics department at Medical College of Wisconsin




MESH terms used to index this publication - Major topics in bold

Antibodies, Monoclonal
Bone Marrow Cells
Cell Separation
Centrifugation
Colony-Forming Units Assay
Diatrizoate
Erythrocytes
Ficoll
Granulocytes
Humans
Lymphocyte Activation
Lymphocyte Culture Test, Mixed
Lymphocyte Depletion
Phytohemagglutinins
T-Lymphocytes
jenkins-FCD Prod-461 7d7c6113fc1a2757d2947d29fae5861c878125ab