Anti-CD4-AF700 Ab For The Detection & Characterization Of Human T-Cells By Flow Cytometry

February 23, 2017

Clinical Research
Albert Einstein College of Medicine
Pre-doctoral researcher

Overall

Quality of Results

Ease-of-Optimization

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Company:

BD Biosciences

Product Name:

Mouse anti-human CD4-Alexa Fluor 700 Ab

Catalog Number:

557922

Using a whole blood staining protocol, we identify and characterize different T-cell sub-populations by flow cytometry using a panel of antibodies that includes BD's anti-CD4-AF700 antibody. We stain for CD4 to differentiate between the two major populations of T-cells, CD4 and CD8 T-cells (Plot CD4 vs. CD8 once CD3+CD45+ cells (total T-cells) have been identified). The antibody described here provides clean separation and reliable staining for all of our studies to date.

Experimental Design and Results Summary

Applications

Flow Cytometry

Sample

Human whole blood (100 ul)

Primary Incubation

Incubate 100 ul whole blood with 5 ul of anti-CD4-AF700 Ab for 20 minutes on ice.

Blocking Agent

Incubate whole blood with 10 ul of human serum for 10 minutes on ice prior to primary incubation.

Secondary Incubation

Add RBC lysis buffer, vortex, and incubate (covered/no light) at room temperature for 15 minutes.

Tertiary Incubation

Add DAPI at 1:10 and acquire data immediately.

Detection

Positive staining can be detected using a flow cytometer (we use an LSR-II).

Results Summary

Following data acquisition, analyses can be performed using flow cytometry software such as FlowJo. Make sure to run single color and FMO controls with each experiment to ensure the utmost accuracy of results. CD4+ cells can be identified once total T-cells (CD3+CD45+) have been gated upon. Plotting against CD8 staining provides good separation.

DOI or PMID #

N/A

Additional Notes

N/A

Related Categories

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Summary

The Good

Easy to optimize, reliable and consistent results (see image which highlights a comparison of our CD4 data against clinically validated testing).

The Bad

Expensive. Make sure to titrate/optimize antibody with your sample type prior to use on precious samples/materials.

The Bottom Line

I recommend this product for use on human whole blood in staining protocols designed to identify, further characterize, or quantify CD4 T-cells. We have never experienced any problems with this product.

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