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Poster
144

Using Impedance-Based Approaches for Measuring Cell-Mediated Cytotoxicity and Antibody-Dependant Cell-Mediated Cytotoxicity (ADCC)

Discussion

The most common method for measuring cell mediated cytotoxicity is the release assay based on the loss of target cell membrane integrity. Within up to 4 hours following effector cell addition in target cell lysis, either the radioactive release from target cells pre-labelled with Chromium-51 or Indium-111 is measured, or the release of naturally occurring substances, such as lactate dehydrogenase (LDH), into the culture medium is assayed. Release of these substances thus serves as an indirect measure of the extent of cell damage due to effector cell-mediated target cell lysis. Alternative endpoint methods also include flow cytometry, enzyme-linked immunosorbent assay-based granzyme measurement, and morphometric analyses by microscopy.

Here we describe an impedance-based real-time label-free method than can automated capture the kinetics of the cell mediated or antibody-dependent cell mediated cytolysis (ADCC) of the target cancer cells. To determine if cell-mediated cytotoxicity, and specifically ADCC, can be investigated using an impedance-based xCELLigence system, the response of tumor cells (as target cells) to natural killer NK) cell activity (as effector cells), in the presence or absence of immunoglobulin G isotype-specific antibody, was measured. Importantly, it is shown that the addition of NK cells in suspension, over a monolayer or adherent tumour cells, does not produce impedance changes, because the NK cells do not come in contact with the electronic sensor. However, the secretion of perforins and granzymes by these non-adherent NK cells does activate caspases inducing tumor cell apoptosis. Dysfunctional and dying tumour cells detach from the sensor electode, reducing the number of viable and adhering cells on the electrode surface.

Overall, our results show that the impedance-base technology is a label-free alternative to the traditional end-point assays. The automated readouts provide direct, sensitive and specific measurement of target cell changes both short term (hrs) and long-term (days). It allows the easy quantification of the cell-mediated cytotoxicity and evaluation of the potencies for specific antibodies.

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