Weak CD223 Staining

August 14, 2015

Immunology
University
Graduate Student

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Company:

eBioscience

Product Name:

Anti-Mouse CD223 (Lag-3) PE

Catalog Number:

12-2231

Info: View Product View Product Specs

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We investigated the expression and upregulation of coinhibitory molecules on antigen-specific T cells as they undergo deletional tolerance.

Experimental Design and Results Summary

Applications

Flow Cytometry

Sample

Murine T cells from lymph nodes and spleen

Primary Incubation

1:100 for 30 min at 4 deg C after cytofix/cytoperm

Blocking Agent

2.4G2 Fc block

Secondary Incubation

NA

Tertiary Incubation

NA

Detection

Canto II flow cytometer

Results Summary

We found weak upregulation of LAG3 on proliferating cells after intracellular staining. Intracellular staining has previously been reported to be required for LAG3 detection (Grosso, 2009). We did not have clear delineation of positive and negative staining using this antibody, but there is definitely some degree of upregulation.

Additional Notes

Intracellular staining using BD cytofix/cytoperm kit

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Summary

The Good

Acceptable LAG-3 staining

The Bad

Probably not staining all LAG-3+ cells

The Bottom Line

Requires isotype control to ensure that positive intracellular staining is real. The staining is not very strong, and it is possible that this is not staining all cells that do express LAG-3. However, what does stain positive is convincing.

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