Treg Detection Kit (CD4/CD25/CD127) ( #130-096-076 #130-096-082 )

Treg Detection Kit (CD4/CD25/CD127)

Overview

The Treg Detection Kit (CD4/CD25/CD127) has been developed as a simple one-reagent solution for the identification of human regulatory T (Treg) cells by flow cytometry. Applications include

Staining and enumeration of CD4⁺CD25⁺CD127^dim/– regulatory T cells in PBMCs, apheresis products, umbilical cord blood, or whole blood
Analysis of Treg cells separated using MACS® Technology

Details

Background information
The kit combines monoclonal antibodies against the standard Treg cell markers CD4, CD25, and CD127 as well as the leukocyte marker CD45 at optimized titers in one vial. The combination of CD4, CD25, and CD127 staining is the current standard method for the identification of live Treg cells. Several publications have reported a strong correlation between the CD25⁺CD127^dim/– phenotype, expression of FoxP3, and suppressive function in CD4⁺ cells.^1,2

Clone	Isotype
CD4 (VIT4)	Mouse IgG2a
CD25 (4E3)	Mouse IgG2b
CD45 (5B1)	Mouse IgG2a
CD127 (MB15-18C9)	Mouse IgG2a

Figure 1

Example of immunofluorescent staining with the Treg Detection Kit (CD4/CD25/CD127) using whole blood and the lyse/no wash protocol with CD45 trigger. A–E depict the gating strategy for the flow cytometric analysis of CD4⁺CD25^high CD127^dim/– cells.

A: An appropriate threshold was defined based on CD45-VioBlue versus side scatter signals, for the exclusion of debris and erythrocytes from the data.

An appropriate threshold was defined based on CD45-VioBlue versus side scatter signals, for the exclusion of debris and erythrocytes from the data.

B: Lymphocytes were gated for their expected morphology using a forward scatter versus side scatter "lymphocyte gate" (P1)

C: Dead cells were excluded by a CD127-PE versus propidium iodide dot plot (P2).

D: CD4⁺ lymphocytes were gated in P3.

CD4<sup>+</sup> lymphocytes were gated in P3.

E: CD25^highCD127^dim/– Treg cells were gated by a CD127-PE versus CD25-APC dot plot.

CD25<sup>high</sup>CD127<sup>dim/–</sup> Treg cells were gated by a CD127-PE versus CD25-APC dot plot.