Development and implementation of natural killer cell simultaneous ADCC and direct killing assay
Maia M.C. Bennett-Boehm,
Anna R. Mahr,
Sean T. Hartwell,
Alexander K. Regan,
Isabelle S. Weber,
Arriana Blackmon,
Cami R. Bisson,
Angela N. Truong,
Bella A. Circo,
Jaden Nienhueser,
Donald R. Rogers,
Nathan Booher,
Nithya Rajagopalan,
Jacob W.S. Martens,
Paul W. Denton
Affiliations
Maia M.C. Bennett-Boehm
Department of Biology, University of Nebraska at Omaha, Omaha, Nebraska, USA; Department of Information Science and Technology, University of Nebraska at Omaha, Omaha, Nebraska, USA
Anna R. Mahr
Department of Biology, University of Nebraska at Omaha, Omaha, Nebraska, USA
Sean T. Hartwell
Department of Biology, University of Nebraska at Omaha, Omaha, Nebraska, USA
Alexander K. Regan
Department of Biology, University of Nebraska at Omaha, Omaha, Nebraska, USA
Isabelle S. Weber
Department of Biology, University of Nebraska at Omaha, Omaha, Nebraska, USA
Arriana Blackmon
Department of Biology, University of Nebraska at Omaha, Omaha, Nebraska, USA
Cami R. Bisson
Department of Biology, University of Nebraska at Omaha, Omaha, Nebraska, USA
Angela N. Truong
Department of Biology, University of Nebraska at Omaha, Omaha, Nebraska, USA
Bella A. Circo
Department of Biology, University of Nebraska at Omaha, Omaha, Nebraska, USA
Jaden Nienhueser
Department of Biology, University of Nebraska at Omaha, Omaha, Nebraska, USA
Donald R. Rogers
Department of Biology, University of Nebraska at Omaha, Omaha, Nebraska, USA
Nathan Booher
Department of Biology, University of Nebraska at Omaha, Omaha, Nebraska, USA
Nithya Rajagopalan
Department of Biology, University of Nebraska at Omaha, Omaha, Nebraska, USA
Jacob W.S. Martens
Department of Biology, University of Nebraska at Omaha, Omaha, Nebraska, USA
Paul W. Denton
Department of Biology, University of Nebraska at Omaha, Omaha, Nebraska, USA; Corresponding author.
Assays to quantify natural killer (NK) cell killing efficacy have traditionally focused on assessing either direct killing or antibody dependent cell-mediated cytotoxicity (ADCC) independently. Due to the probability that immunotherapeutic interventions affect NK cell-mediated direct killing and NK cell-mediated ADCC differently, we developed an assay with the capacity to measure NK cell-mediated direct killing and ADCC simultaneously with cells from the same human donor. Specifically, this design allows for a single NK cell population to be split into several experimental conditions (e.g., direct killing, ADCC), thus controlling for potential confounders associated with human-to-human variation when assessing immunotherapy impacts. Our Natural Killer cell Simultaneous ADCC and Direct Killing Assay (NK-SADKA) allows researchers to reproducibly quantify both direct killing and ADCC by human NK cells. Furthermore, this optimized experimental design allows for concurrent analysis of the NK cells via flow cytometric immunophenotyping of NK cell populations which will facilitate the identification of relationships between NK cell phenotype and the subsequent killing potential. This assay will be valuable for assessing the broader impact(s) of immunotherapy strategies on both modes of NK cell killing.
