Good BD Biosciences Antibody for CD4 detection on Lymphocytes

July 02, 2019

BlueskyReddit
Division of BioMedical Sciences, Faculty of Medicine
Memorial University of Newfoundland
Research Assistant

Overall

Quality of Results

Ease-of-Optimization

What do these ratings mean?
Write a Review

Company:

BD Biosciences

Product Name:

BV650 Mouse Anti-Human CD4

Catalog Number:

563737

Clone Number:

L200

Browse Similar Products: CD4 Flow Cytometry Antibodies

Our lab routinely uses flow cytometry to perform immunophenotyping studies of primary cells in different disease states. Our goal in using this product was to identify CD4+ T cells in human peripheral blood mononuclear cells as part of a larger immunophenotyping panel that we are developing. This particular antibody clone was chosen because it is available with several different conjugated fluorophores. More specifically, the BV650-conjugated version was chosen because it is compatible with our flow cytometer and other antibodies in our flow cytometry antibody panel.

Experimental Design and Results Summary

Applications

Flow Cytometry

Sample

Human PBMC

Primary Incubation

BV650-conjugated mouse anti-human CD4 antibody (BD 563737), incubated with approximately 5x10^5 cells for 30 minutes on ice at a dilution of 1:20 in a total volume of 100ul PBS+2%FCS, 1mM EDTA (5ul antibody).

Blocking Agent

None

Secondary Incubation

None

Tertiary Incubation

None

Detection

Beckman Coulter CytoFLEX flow cytometer

Results Summary

This antibody worked well in flow cytometric analysis of human PBMC. CD4 staining was very bright as expected since BV650 is classified as a 'very bright' compared to other fluorophores according to the manufacturer. CD4+ cells were well separated from CD4- cells. As expected, significant spectral spillover was observed in the APC and Alexa Fluor 700 detectors (not shown) since BV650 is a tandem dye with the acceptor fluorophore being excited with the 640nm laser.

DOI or PMID #

N/A

Additional Notes

For flow cytometric analysis, gating was performed on the lymphocyte population in the FSC vs SSC plot.

Related Categories

Image Gallery

Summary

The Good

Bright staining. Does not need much optimization other than initial titrating. Clone is available in several formats allowing you to switch fluorophores when required.

The Bad

BV650 is a tandem dye, so extra precautions are necessary (extra light sensitive, control for lot to lot variations, etc). Significant spillover into the APC and Alexa Fluor 700 detectors with our flow cytometer configuration.

The Bottom Line

Overall, this is an easy-to-use antibody that results in very bright staining of CD4+ cells. Depending on the application, you might want to use this clone conjugated to a different fluorophore if the spillover into the APC and Alexa Fluor 700 detectors is problematic for your assay design / instrument configuration or if you tend to avoid the use of tandem dyes entirely.

Share your experience with other scientists. Write a Review! »

Join the discussion