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CellXVivo Human Treg Cell Differentiation Kit

For the differentiation of human regulatory T (Treg) cells from a preparation of naïve CD4+ T cells.
Differentiation of Human CD4+ T Cells into Treg Cells Confirmed by FoxP3 and CD25 Expression.
(2)
CellXVivo Human Treg Cell Differentiation Kit
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CDK006
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Assay Procedure

Refer to the product datasheet for complete product details.

Briefly, mature human CD4+ T cells can be differentiated into Tregs using the following procedure:

  • Coat a plate with Mouse Anti-Human CD3 Antibody
  • Isolate naïve CD4+ T cells from Human PBMCs
  • Culture naïve CD4+ T cells in Human Treg Differentiation Media for 5 days
  • Verify differentiation into Treg cells by flow cytometry
 

 

Reagents Provided

Reagents Supplied in the CellXVivo™ Human T (Treg) Cell Differentiation Kit (Catalog # CDK006):

  • Mouse Anti-Human CD3 Antibody
  • Human Treg Reagent 1
  • Human Treg Reagent 2
  • Human Treg Reagent 3
  • Human Treg Reagent 4
  • Reconstitution Buffer 1
  • Reconstitution Buffer 2
  • Wash Buffer (20X)

 

Other Supplies Required

Reagents

  • Ficoll-Hypaque™
  • MagCellect™ Human Naïve CD4+ T Cell Isolation Kit (R&D Systems, Catalog # MAGH115, or equivalent)
  • X-VIVOTM15 Chemically Defined, Serum-free Hematopoietic Cell Medium (Lonza, or equivalent)

Equipment

  • Tissue culture flasks and/or plates
  • Sterile deionized water
  • Microscope
  • Hemocytometer
  • 37 °C and 5% CO2 incubator
  • Centrifuge
  • Pipettes and pipette tips

 

Procedure Overview

Coat wells of a 24-well plate with Mouse Anti-Human CD3 Antibody.

Coat wells of a 24-well plate

Isolate PBMCs from human blood.

Isolate PBMCs from human blood

Isolate human naïve CD4+ T cells from PBMCs (e.g., using magnetic cell selection).

Isolate T Cells from PBMCs

Perform a cell count.

Perform a cell count

Suspend 1 - 2 x 105 naïve CD4+ T cells/mL in Human Treg Differentiation Media.

Culture the cells on plates pre-coated with CD3 antibody for 5 days.

Suspend cells in media

Verify Treg cell differentiation by analyzing marker expression using flow cytometry.

Mature Treg cells are now ready for further downstream applications.

Verify Treg cell differentiation

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Product Datasheets

Certificate of Analysis

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Note: Certificate of Analysis not available for kit components.

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