Rapid flow cytometry based cytotoxicity assay for evaluation of NK cell function

Area of Science:

• Immunology
• Cellular Biology
• Diagnostic Assays

Background:

• Natural killer (NK) cell cytotoxicity assessment is crucial for research and disease diagnosis.
• Current methods, like chromium release or flow cytometry, often require large blood volumes and PBMC isolation, posing challenges for pediatric or cytopenic patients.

Purpose of the Study:

• To develop and validate a novel, rapid flow cytometric assay for NK cell activity using whole blood.
• To reduce blood volume requirements and processing time compared to traditional PBMC-based assays.

Main Methods:

• A dual fluorescent staining flow cytometry assay was designed using K562 target cells labeled with DIOC18 and counterstained with 7-AAD.
• NK cell cytotoxicity was measured by incubating labeled target cells with whole blood and analyzing dead target cells via flow cytometry.
• The whole blood assay was compared against the PBMC-based assay using clinical samples and validated in patients with familial hemophagocytic lymphohistiocytosis.

Main Results:

• The whole blood NK cell cytotoxicity assay demonstrated no significant difference in lytic activity or reproducibility compared to the PBMC assay.
• The whole blood assay required significantly less blood volume and processing time.
• The assay successfully identified reduced NK cell activity in patients with familial hemophagocytic lymphohistiocytosis.

Conclusions:

• A rapid, sensitive, and reproducible whole blood flow cytometry assay for NK cell activity has been developed.
• This assay simplifies NK cell function testing, requiring less blood and processing time, making it ideal for clinical evaluation, especially in vulnerable patient populations.