Clearly Labels Foxp3 Expression Among CD4+ T Cells

January 13, 2017

Immunology
Duke University
Predoctoral

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Company:

eBioscience

Product Name:

Anti-Mouse/Rat Foxp3 PE-Cyanine5

Catalog Number:

15-5773-82

The goal of this research was to identify CD4+ regulatory T cells in the lymph nodes and spleens of tumor-bearing mice. I selected this product because Ebioscience is the only company that produces a high-quality Foxp3-binding antibody, and this color fit in well with my staining panel.

Experimental Design and Results Summary

Applications

Flow Cytometry

Sample

Mouse Lymph Node and Splenic Leukocytes

Primary Incubation

Wash cells once in FACS buffer (PBS plus 1% BSA or serum). Block Fc receptors and stain for viability in PBS for 20 minutes on ice.

Blocking Agent

Fc block

Secondary Incubation

Wash cells once in FACS buffer. Stain cell surface molecules with antibody (1:500 dilution, 100 uL per million cells) for 20 minutes on ice. Wash cells once in FACS buffer. Permeabilize cells using Ebioscience's Foxp3/Transcription Factor Buffer Staining Set (cat. 00-5523) according to manufacturer's instructions.

Tertiary Incubation

Stain with isotype control or Foxp3 antibody (1:100 dilution, 100 uL per million cells) for 30-60 minutes on ice. Wash cells and resuspend cells in 100-250 uL PBS containing 1.5% paraformaldehyde.

Detection

Flow Cytometer

Results Summary

Regulatory T cells expressing Foxp3 were easily distinguished in the lymph nodes (left panel) and spleens (left panel) of tumor-bearing mice.

DOI or PMID #

N/A

Additional Notes

The Ebioscience Foxp3/Transcription Factor Buffer Staining Set (cat. 00-5523) must be used to permeabilized cells appropriately for Foxp3 visualization.

Related Categories

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Summary

The Good

Brightly labels Foxp3 in CD4 T cells.

The Bad

It must be used in conjunction with Ebioscience's transcription factor buffer staining set.

The Bottom Line

Ebioscience makes the best Foxp3-directed antibody for flow cytometry, so this is a safe way to go, if the price tag of the antibody and buffer staining set aren't too high.

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