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A Flow Cytometry-Based Cytotoxicity Assay for the Assessment of Human NK Cell Activity
Published on: August 9, 2017
The correlation between the percent of CD3- CD56+ cells and NK precursor function
Ahmad Gharehbaghian1, Craig Donaldson, John Newman
1The Research Centre of Iranian Blood Transfusion Organisation, Hemmat express way, Next to the Milad tower, P.O.Box: 14665-1157, Tehran, Iran. gharehbaghian@ibto.ir
Iranian Journal of Allergy, Asthma, and Immunology
|January 24, 2007
Summary
Assessing natural killer (NK) cell function often uses CD56 as a marker. This study found that CD56 alone is inadequate for identifying functional NK cells, as there was no correlation with NK precursor frequency.
Area of Science:
- Immunology
- Cell Biology
Background:
- Natural killer (NK) cells are crucial for immune surveillance, recognizing and eliminating target cells without prior sensitization.
- NK cell number and function are key indicators for monitoring immune status, treatment efficacy, and disease progression, including malignancy and metastases.
- Human NK cells are typically identified by surface markers CD16 and CD56, with CD56 widely used as a primary hallmark.
Purpose of the Study:
- To investigate the unique subsets of peripheral blood mononuclear cells (PBMC), specifically %CD3-CD56+ cells, using flow cytometry.
- To determine if there is a correlation between these identified PBMC subsets and the functionally mature progeny of natural killer cell precursors (NKp) after a five-day culture period.
- To evaluate the adequacy of CD56 as a sole marker for functional NK cells.
Main Methods:
- Peripheral blood samples from 120 Caucasian patients were analyzed.
- NK precursor frequency (NKpf) was measured using limiting dilution analysis (LDA) and a specific NKpf assay.
- Functional maturity of NK cells was assessed by their cytotoxic activity against the K562 cell line using Europium (Eu) release assays and time-resolved fluorometry.
- Immunophenotyping by flow cytometry was performed using anti-CD3 and anti-CD56 antibodies to enumerate NK cells.
Main Results:
- Flow cytometry analysis revealed no correlation between the natural killer precursor frequency (NKpf) and the percentage of CD3-CD56+ cells after five days of culture.
- This finding indicates that CD56 expression alone is insufficient to reliably identify functionally mature NK cells.
- The study highlights a discrepancy between NK cell surface marker expression and functional capacity.
Conclusions:
- The commonly used CD56 marker is inadequate as a unique identifier for functional natural killer cells.
- Assessing NK cell function requires methods beyond simple surface marker enumeration, such as functional assays measuring cytotoxicity.
- Further research is needed to identify reliable markers that correlate with NK cell precursor maturation and functional competence.
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