Identifying Leukocytes In Whole Blood By Flow Cytometry Using Anti-CD45-APC

February 16, 2017

Clinical Research
Albert Einstein College of Medicine
Pre-doctoral researcher

Overall

Quality of Results

Ease-of-Optimization

What do these ratings mean?
Write a Review

Company:

BD Pharmingen

Product Name:

Mouse anti-human CD-45-APC

Catalog Number:

555485

We use a whole blood staining protocol to identify different lymphocyte and monocyte populations in human samples. BD's anti-human-CD45-APC antibody allows us to identify lymphocytes, monocytes, and granulocytes from not only one another but also from platelets and dead cells/debris (when plotted vs. SSC). The antibody is consistent and provides very clean separation of our cell populations of interest.

Experimental Design and Results Summary

Applications

Flow Cytometry

Sample

Human whole blood (100 ul)

Primary Incubation

Incubate 100 ul whole blood with 20 ul of anti-CD45-APC 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.

DOI or PMID #

N/A

Additional Notes

N/A

Related Categories

Image Gallery

Summary

The Good

Relatively inexpensive, easy to optimize, reliable and consistent results.

The Bad

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

The Bottom Line

I highly recommend this antibody for use in whole blood staining protocols for the detection of human leukocytes -- including monocytes, lymphocytes, and granulocytes -- by flow cytometry. Once optimized on your cell population(s) of interest, the protocol is incredibly easy to follow and the results are consistent/reliabe.

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

Join the discussion