RT Book, Section T1 Assessing changes in human natural killer cell metabolism using the seahorse extracellular flux analyzer. A1 Traba, Javier A1 Antón Hurtado, Olga María AB Natural killer (NK) cells are cytotoxic cells that mediate anti-tumor and anti-viral immunity. The response of NK cells to different cytokines and stimuli may involve cell survival, proliferation, and changes in their cytotoxic function. These responses will be supported by changes in cellular metabolism. Therefore, changes in NK metabolic parameters could somehow predict changes in NK cell function and cytotoxicity. In this chapter, we describe a protocol to measure NK cell metabolism in primary human NK cells by using an extracellular flux analyzer. This machine measures pH and oxygen changes in the medium and allows the study of NK cell glycolysis and mitochondrial respiration in real time with a small number of cells. SN 978-1-0716-2159-2 SN /978-1-0716-2160-8 SN 1064-3745 SN 1940-6029 YR 2022 FD 2022 LK https://hdl.handle.net/20.500.14352/100097 UL https://hdl.handle.net/20.500.14352/100097 LA eng NO Traba, J., Antón, O.M. (2022). Assessing Changes in Human Natural Killer Cell Metabolism Using the Seahorse Extracellular Flux Analyzer. In: Shimasaki, N. (eds) Natural Killer (NK) Cells. Methods in Molecular Biology, vol 2463. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2160-8_12 NO We thank Michael N. Sack (National Heart, Lung and Blood Institute, NIH) and Thomas A. Waldmann (National Cancer Institute, NIH) for continuous support and discussion. This study was supported by grants RYC2018-026050-I and PID2019- 105665RA-I00 of MICINN (Spain) to J.T. This study was also supported by the Intramural Research Programs of the National Institutes of Health, National Cancer Institute, and National Heart, Lung and Blood Institute. NO Ministerio de Ciencia e Innovación (España) DS Docta Complutense RD 4 abr 2025