FITC-Conjugated Monoclonal Mouse Anti-Human CD3 Antibody (for Flow Cytometry) from BioLegend (Clone SK7)

May 19, 2016

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

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Company:

BioLegend

Product Name:

FITC anti-human CD3 Antibody

Catalog Number:

344804

We used this antibody to identify CD3+ T cells from human peripheral blood mononuclear cells (PBMC) by flow cytometry. The antibody performed well in that there was a clear separation of CD3-positive and -negative cells.

Experimental Design and Results Summary

Applications

Flow Cytometry

Sample

Human PBMC isolated from blood by standard Ficoll density gradient separation

Primary Incubation

FITC anti-human CD3 antibody (BioLegend 344804), incubated with approximately 1x10^6 cells for 1 hour on ice at 1:20 dilution in a total volume of 100ul PBS+2% FCS, 1mM EDTA, 0.1% sodium azide.

Blocking Agent

N/A

Secondary Incubation

N/A

Tertiary Incubation

N/A

Detection

FACSCalibur (Becton Dickinson)

Results Summary

This antibody worked well for FACS staining of human PBMC. CD3-expressing cells were bright and well separated from negative cells, making the distinction between the two populations fast and simple. Some may want to titrate this antibody so that it is used at a higher dilution than 1:20 so that negative cells exhibit similar fluorescence as unlabelled cells, but this is not necessary in my opinion. The epitope recognized by this antibody is extracellular, thus fixing and permeabilization of cell is not required resulting in a quick and simple procedure for staining cells.

Additional Notes

For flow cytometric analysis, gating was performed on the lymphocyte population. Monocytes were negative (not shown).

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Summary

The Good

Bright staining. Directly conjugated and recognizes extracellular epitope = quick and simple staining procedure. Affordable pricing.

The Bad

Nothing really

The Bottom Line

This antibody is recommended for staining of CD3+ cells in human PBMC by flow cytometry. It performs comparably to other well established monoclonal anti-human CD3 antibodies and is worth consideration.

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