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J Immunol Methods. 2013 Oct 31;396(1-2):152-6. doi: 10.1016/j.jim.2013.07.004. Epub 2013 Jul 31.

Efficient depletion of chronic lymphocytic leukemia B cells using serial rounds of immunomagnetic depletion.

Author information

1
Malaghan Institute of Medical Research, Wellington, New Zealand; Wellington Blood & Cancer Centre, Capital & Coast District Health Board, Wellington, New Zealand; Department of Pathology and Molecular Medicine, University of Otago Wellington, Wellington, New Zealand. Electronic address: rweinkove@malaghan.org.nz.

Abstract

Functional studies of cellular immunity in patients with leukemia often require separation of leukemic cells from other peripheral blood mononuclear cells (PBMCs). This can pose a challenge when the number of leukemic cells is very high, such as in untreated patients with chronic lymphocytic leukemia (CLL). We found that when leukemia cell frequency was very high, anti-CD19 coated immunomagnetic beads did not thoroughly deplete B cells when used according to manufacturer's instructions. In this study, we depleted leukemic B cells using a modified protocol comprising serial rounds of depletion using immunomagnetic beads at reduced bead to cell ratios. This resulted in more effective B cell depletion with the use of fewer immunomagnetic beads, and without affecting viability or yield of non-B cells. CD19- PBMC subsets were retained, and serial depletion rounds did not activate T cells and monocytes. The positively isolated CLL cells were of high purity and were available for downstream analysis. This is a convenient and cost-effective method to enable in vitro analysis of immunocompetent cells from patients with leukemia.

KEYWORDS:

B cell antigen CD19; B-cells; Chronic lymphocytic leukemia; Immunomagnetic cell separation

PMID:
23911312
DOI:
10.1016/j.jim.2013.07.004
[Indexed for MEDLINE]

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