Staining Mouse Antigen Presenting Cells With PE Anti-Mac-2

Pediatrics
UW-Madison
Postdoc

Overall

Quality of Results

Ease-of-Optimization

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Company:

BioLegend

Product Name:

PE anti-mouse/human Mac-2 (Galectin-3) Antibody

Catalog Number:

125405

Mac2, also known as Galectin-3, is a pattern recognition receptor on various leukocyte populations. I titrated the anti-mouse Mac2 antibody on cells from the spleen or lymph nodes from naïve mice. Resident DCs (MHC class II+, CD11c+, CD11b-, F4/80-) in these lymphatic tissues had one of the largest proportion of Mac2+ cells in naïve mice. I titrated the antibody in the concentration range suggested by BioLegend finding that 1:200 dilution of the antibody stained better than 1:100 and 1:50.

Experimental Design and Results Summary

Applications

Flow Cytometry

Sample

Splenocytes or cells from skin-draining lymph nodes from naive C57B/6 mice

Primary Incubation

Titrations (as shown) for 30 minutes at 4C along with antibodies against other markers for gating.

Blocking Agent

Buffer contained BSA and FcBlock

Secondary Incubation

Cells were fixed with 2% paraformaldehyde

Tertiary Incubation

None

Detection

Flow cytometry

Results Summary

I show gated DCs from the spleen and pooled skin-draining LNs from naïve C57B/6 mice (MHC class II+, CD11c+, CD11b-, F4/80-). The same sample data is shown after staining with different concentrations of anti-Mac2-PE. Around 5% of these DCs were positive for Mac-2. While the staining profile was not incredibly different among the titrations, the 1:200 dilution was the best.

DOI or PMID #

N/A

Additional Notes

I have been using this product at 1:200 on antigen-presenting cells since this titration experiment with good data.

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Summary

The Good

Stains well

The Bad

None

The Bottom Line

This product works well. I have been using it assays with success.

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