Abstracts (complete list) - Wissenschaft Online

Carina Conrads, Ramona Siemer, Mario Assenmacher, Claudia Niemand
Quality assessment of enriched CD4+CD25+ regulatory T cells
Many publications during the last years have reported naturally occurring CD4+CD25+
regulatory T cells to play an important role in autoimmunity, transplantation tolerance
and tumor immunity. Intracellular FoxP3 staining is an important method for the
analysis of regulatory T cells. In addition functional capacity of regulatory T cells is often
assessed by in vitro suppression assays.
We have developed a ready-to-use, well-defined, and robust method using
MACSiBeadTM Particles for in vitro suppression assays. MACSiBeadTM Particles loaded
with CD2, CD3 and CD28 antibodies act as artificial APCs and are simply added in a 1:1
bead to cell ratio to the cell culture.
Human CD4+CD25+ regulatory T cells were isolated from peripheral white blood cells
using the MACS technology (n=9) with a mean purity of 87.8% (range 76.5-93.6%).
More than 95% of isolated cells were CD4+, of which 78.5% were positive for FoxP3
(range 63.8-88.5%).
Isolated CD4+CD25+ regulatory T cells and autologous CD4+CD25- responder T cells
were polyclonally stimulated with CD2, CD3 and CD28 antibody loaded MACSiBeadTM
Particles (“Treg Suppression Inspector”) for 4 days and proliferation was analyzed by
3H-thymidine incorporation for 16 hours. Isolated regulatory CD4+CD25+ T cells
showed a mean suppression rate of 67.6% (range 38-92%) at a 1:1 ratio of regulatory
T cells to responder T cells.
We show that quality assessment of enriched CD4+CD25+ regulatory T cells is possible
by staining for FoxP3 and using MACSiBeadTM Particles.