CD4
⁺
CD25
⁺
CD127
^dim/-
Regulatory T Cell Isolation Kit II
, human

Name: CD4+ CD25+ CD127dim/- Regulatory T Cell Isolation Kit II, human
Availability: InStock

Product overview

The CD4

⁺

CD25

⁺

CD127

^dim/–

Regulatory T Cell Isolation Kit II was developed for the isolation of CD4

⁺

CD25

⁺

CD127

^dim/–

Treg cells from human PBMCs. The isolation is performed in a two-step procedure.

Background information

CD127, the α-chain of the IL-7 receptor, is expressed on the majority of mature T cells and plays an important role in their proliferation and differentiation. However, on regulatory T cells CD127 is absent and its expression inversely correlates with FoxP3 expression. Thus, CD127 can be used as an additional marker to discriminate between human regulatory and activated T cells.

Detailed separation procedure

First, the isolation of CD4

⁺

CD25

⁺

CD127

^dim/–

regulatory T cells was performed with a cocktail of biotinylated antibodies and Anti-Biotin MicroBeads for the depletion of non-CD4

⁺

and CD127

^high

cells. In the second step, the flow-through fraction of pre-enriched CD4

⁺

CD127

^dim/–

T cells is labeled with CD25 MicroBeads for subsequent positive selection of CD4

⁺

CD25

⁺

CD127

^dim/–

regulatory T cells.

Downstream applications

Isolation of CD4

⁺

CD25

⁺

CD127

^dim/–

regulatory T cells from human PBMCs for further phenotypical or functional characterization.

Columns

For the first magnetic separation (depletion): LD or autoMACS

^®

Columns. For the second magnetic separation (positive selection): MS or autoMACS Columns.

Product data and images

Figure 1

CD4

⁺

CD25

⁺

CD127

^dim/–

regulatory T cells were isolated from human PBMCs by using the CD4

⁺

CD25

⁺

CD127

^dim/–

Regulatory T Cell Isolation Kit II, an LD and two MS Columns, a MidiMACS™ Separator and a MiniMACS™ Separator. The cells were fluorescently stained with CD4-FITC, CD25-APC, and CD127-PE, or CD4-FITC, CD127-PE, and Anti-FoxP3-APC and analyzed by flow cytometry using the MACSQuant® Analyzer. Gating was performed according to CD4 expression, except for the CD25-APC vs. CD4-FITC dot plot.

PBMCs before separation


Pre-enriched CD4⁺ CD127^dim/- cells


Isolated CD4⁺ CD25⁺ CD127^dim/- regulatory T cells

Scientific resources

Selected references
1. Seddiki, N. et al. (2006) Expression of interleukin (IL)-2 and IL-7 receptors discriminates between human regulatory and activated T cells. J. Exp. Med. 203: 1693-1700
2. Liu, W. et al. (2006)
  CD127 expression inversely correlates with FoxP3 and suppressive function of human CD4
  ⁺
  T reg cells.
  J. Exp. Med. 203: 1701-1711
3. Stockis et al. (2009) Membrane protein GARP is a receptor for latent TGF-beta on the surface of activated human Treg. Eur. J. Immunol. 39: 3315-3322
Brochures and posters

Regulatory T cell research
Customer reports and application notes

Human CD4
⁺
CD25
⁺
CD127
^dim/-
regulatory T cell isolation,
in vitro
expansion and analysis - Workflow protocol

Product options: 1

130-094-775