A Great Alternative to Mouse Antibodies to Make Your Immunofluorescence Life Easier

July 06, 2016

Institute of Inflammation and Ageing
University of Birmingham
Research Associate

Overall

Quality of Results

Ease-of-Optimization

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Company:

Abcam

Product Name:

Anti-CD3 antibody

Catalog Number:

ab16669

I needed to be able to fit CD3 into my existing antibody panel for immunoflourescence within my tissue. As I study human tissue the antibodies that I was already using were IgG1, 2a, 2b and rat. As I couldn't use traditional CD3 clones (UCHT1 and OKT3), I thought I'd try this. It works and really helps for fitting in that fourth colour.

Experimental Design and Results Summary

Applications

Immunofluorescence

Sample

Human tonsil and synovium

Primary Incubation

1 in 50, 1 hour at RT

Blocking Agent

Goat serum

Secondary Incubation

Goat anti rabbit alexa flour 488, 1/100, 45 mins, RT.

Tertiary Incubation

N/a (but I do use Hoechst for my nuclei)

Detection

Confocal microscopy (but fine with a fluorescence microscope)

Results Summary

Good staining within the T zone of tonsil and in my synovial tissues. I also had the same quality staining with goat anti rabbit Alexa Flour 546 as the secondary (1/300). Abcam are now testing all their antibodies with Cripr/Cas9 knock outs so That's very reassuring.Image shows the T cell and B cell interface within a human tonsil. B cells in blue with CD20, T cells in green.

Additional Notes

None

Related Categories

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Summary

The Good

A great way to get your generic T cell marker in to your complicated antibody panels.

The Bad

Check your secondaries aren't raised in rabbit!

The Bottom Line

If your PI/collaborator asks "can you just look for this" and you know that it's going to mean a major reshuffle in the staining you want to do this is an excellent way of getting out of a jam. And for staining for human CD3 in general.

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