IgM Alexa Fluor 647

September 03, 2015

Microbiology and Immunology
University of Miami
Post-doctorall fellow

Overall

Quality of Results

Ease-of-Optimization

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Company:

Jackson ImmunoResearch

Product Name:

Alexa Fluor® 647 AffiniPure Fab Fragment Goat Anti-Mouse IgM, µ Chain Specific

Catalog Number:

115-607-020

Info: View Product View Product Specs

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Our lab is concerned with studying the transcriptome and signaling changes that occur during transitional B-cell development. This is a major checkpoint in peripheral tolerance where autoreactive B-cell clones are purged. IgM AF647 was used to isolate various B-cell subsets. This antibody not only provides exceptional separation but is unlikely to cause any BCR cross linking.

Experimental Design and Results Summary

Applications

Flow Cytometry

Sample

C57BL/6 splenocytes

Primary Incubation

15-20min in FACS buffer 1:400

Blocking Agent

Fc block 5min prior

Secondary Incubation

NA

Tertiary Incubation

NA

Detection

Flow cytometry

Results Summary

This antibody provides the most separation of any IgM antibody tested in our lab. It is a very strong antibody so dilution must be optimized. Lower dilutions than 1:400 are probably okay. Can be used for fixed and non-fixed staining.

Additional Notes

Aliquots can be frozen -80 and used later

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Summary

The Good

IgM AF647 provides exceptional separation and can be used in a range of flow applications

The Bad

Antibody optimization due to strength of fluorochrome

The Bottom Line

I would highly recommend this antibody to those doing regular staining of B-cells for either flow cytometry or FACS sorting.

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