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March 09, 2017

Antibody dependent cell-mediated cytotoxicity (ADCC)

Background:

The aim of this assay is to assess the effects of test antibodies on the cytotoxic activity of NK cells.

NK cells express the FcγIII receptor (CD16). This receptor can bind to the Fc part of the antibody (Ab) which has bound to the specific antigen on the target cell. Cross-linking of the Fc receptors induces the NK cells to release perforin and granzymes, which causes the lysis of the target cell.

Method:

The assay is carried out by co-culturing purified NK cells as effector cells in the presence of the test antibody Ab (at different concentrations) with fluorescently labeled (CFSE-stained) target cells. The number of target cells killed by NK cells is determined by flow cytometry using a fluorescent dead-cell dye (7-AAD).

Outcome:

Dead cells are stained after the incubation period with the dead-cell dye 7-AAD and samples are subjected to flow cytometry analysis. CFSE+/7-AAD+, double positive cell population, represents the dead cells indicating the cytotoxic effect of NK cells.

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