Miltenyi, 130-092-614, Double-negative T Cell Isolation Kit, human

Background information
The down-regulation of immune responses by regulatory T cells plays a key role in the induction of tolerance. Recently, it was reported that TCRα/β+ CD4–CD8– double-negative T cells also have the ability to specifically down-regulate immune responses towards allo-antigens both in humans and mice. Double-negative T cells are found in lymphoid as well as in non-lymphoid tissues. They constitute about 1–3% of peripheral CD3+ cells. It has been described that double-negative T cells can take up allo-MHC peptide complexes from antigen-presenting cells.

Detailed separation procedure
The isolation is performed in a two-step procedure. To remove all unwanted TCRα/β+ cells, firstly the CD4+, CD8+, and CD56+ cells are indirectly magnetically labeled and depleted. Labeling with Anti-TCRα/β-PE is performed simultaneously. In the second step, the flow-through fraction with the pre-enriched double-negative T cells is incubated with Anti-PE MicroBeads, and target cells are isolated by positive selection.

Columns
For the first magnetic separation (depletion): LD or autoMACS® Columns. For the second magnetic separation (positive selection): MS or autoMACS Columns.

Overview
The Double-negative T Cell Isolation Kit was developed for the isolation of double-negative T cells (CD4–CD8–CD56–CD3+TCRα/β+ cells) from PBMCs by sequential sorting. The isolation is performed in a two-step procedure.